ABCMdb

ABCC7 p.Gly91Arg

ClinVar:	c.271G>A , p.Gly91Arg D , Pathogenic
CF databases:	c.271G>A , p.Gly91Arg (CFTR1) D , This mutation was observed through DGGE screening and direct DNA sequencing. The substitution is a G->A at nucleotide position 403. It changes a glycine residue for an arginine G91R. The haplotype bearing the mutation is a C haplotype. The CF child has a [delta]F508 on th other chromosome. He is 9 years old and pancreatic sufficient. The substitution was observed once on 87 non-[delta]F508 chromosomes and non oberved on 70 [delta]F508 chromosomes.
Predicted by SNAP2:	A: D (75%), C: D (85%), D: D (91%), E: D (91%), F: D (95%), H: D (95%), I: D (95%), K: D (95%), L: D (91%), M: D (95%), N: D (91%), P: D (95%), Q: D (91%), R: D (59%), S: D (85%), T: D (85%), V: D (85%), W: D (95%), Y: D (95%),
Predicted by PROVEAN:	A: N, C: D, D: N, E: N, F: D, H: N, I: D, K: D, L: D, M: D, N: N, P: D, Q: N, R: D, S: N, T: N, V: D, W: D, Y: N,

[switch to compact view]
Comments [show]

None has been submitted yet.

Publications
[hide] Insight in eukaryotic ABC transporter function by ... FEBS Lett. 2006 Feb 13;580(4):1064-84. Epub 2006 Jan 19. Frelet A, Klein M
Insight in eukaryotic ABC transporter function by mutation analysis.
FEBS Lett. 2006 Feb 13;580(4):1064-84. Epub 2006 Jan 19., 2006-02-13 [PMID:16442101]

Abstract [show]
With regard to structure-function relations of ATP-binding cassette (ABC) transporters several intriguing questions are in the spotlight of active research: Why do functional ABC transporters possess two ATP binding and hydrolysis domains together with two ABC signatures and to what extent are the individual nucleotide-binding domains independent or interacting? Where is the substrate-binding site and how is ATP hydrolysis functionally coupled to the transport process itself? Although much progress has been made in the elucidation of the three-dimensional structures of ABC transporters in the last years by several crystallographic studies including novel models for the nucleotide hydrolysis and translocation catalysis, site-directed mutagenesis as well as the identification of natural mutations is still a major tool to evaluate effects of individual amino acids on the overall function of ABC transporters. Apart from alterations in characteristic sequence such as Walker A, Walker B and the ABC signature other parts of ABC proteins were subject to detailed mutagenesis studies including the substrate-binding site or the regulatory domain of CFTR. In this review, we will give a detailed overview of the mutation analysis reported for selected ABC transporters of the ABCB and ABCC subfamilies, namely HsCFTR/ABCC7, HsSUR/ABCC8,9, HsMRP1/ABCC1, HsMRP2/ABCC2, ScYCF1 and P-glycoprotein (Pgp)/MDR1/ABCB1 and their effects on the function of each protein.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
359 G85E and G91R affected folding by insertion of a charged residue within the plane of the bilayer [169]. Login to comment

[hide] Defects in processing and trafficking of the cysti... Kidney Int. 2000 Mar;57(3):825-31. Skach WR
Defects in processing and trafficking of the cystic fibrosis transmembrane conductance regulator.
Kidney Int. 2000 Mar;57(3):825-31., [PMID:10720935]

Abstract [show]
Cystic fibrosis (CF) is caused by inherited mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-regulated chloride channel expressed in epithelial tissues. Most mutations in CF patients result in rapid intracellular degradation of the CFTR protein. While this defect is thought to result from abnormal protein folding, it is unclear how mutant and wild-type (WT) proteins differ in structure, how the cell is able to distinguish these differences, and how the fate of the mutant protein is determined. By examining the initial steps of CFTR assembly into the endoplasmic reticulum (ER) membrane, it has recently been shown that CFTR utilizes two redundant translocation pathways to direct N-terminus folding events. Mutations that block one pathway therefore do not alter transmembrane topology, but rather appear to disrupt intracellular trafficking through perturbations in higher order tertiary structure. These studies suggest that cellular quality control machinery acts at least in part, by monitoring proper interactions between CFTR subdomains. The end result of this process is the conversion of misfolded CFTR into a membrane-bound, polyubiquitinated complex. This complex recruits cytosolic degradation machinery to the endoplasmic reticulum membrane where CFTR is degraded as it is extracted from the lipid bilayer. Understanding how cellular machinery mediates this process will be an important step in designing strategies to modify protein folding and degradation in CF and related ion channelopathies.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
52 The post-translational pathway is utilized by most (Ͼ60%) of WT chains and essentially all G85E and G91R mutant chains. Login to comment
76 While it is often tempting Two CF mutations, G85E and G91R, each introduce to view the acquisition of protein function as a criteria an additional charged residue within the hydrophobic for "normal" folding, in the case of CFTR this is not core of TM1. Login to comment
79 This suggested that G85E and G91R CFTR mutants [35]. Login to comment

[hide] Genotype analysis and phenotypic manifestations of... Chest. 2000 Dec;118(6):1591-7. Desmarquest P, Feldmann D, Tamalat A, Boule M, Fauroux B, Tournier G, Clement A
Genotype analysis and phenotypic manifestations of children with intermediate sweat chloride test results.
Chest. 2000 Dec;118(6):1591-7., [PMID:11115444]

Abstract [show]
STUDY OBJECTIVES: Cystic fibrosis (CF) is one of the most common inherited diseases among whites. Since the cloning of the CF transmembrane conductance regulator (CFTR) gene, a number of studies have focused on associations between the genotype and phenotype in CF. This had led to the progressive identification of new groups of patients, including those who have mild lung disease and those who have normal sweat chloride values (< 60 mEq/L). The aim of the present work was to provide information on the genotype and the phenotypic characteristics of children with intermediate-range sweat chloride test results. PATIENTS AND RESULTS: We focused on children referred to the pulmonary department for various types of pulmonary disease and who had several sweat chloride test results with median values in the range of 40 to 60 mEq/L. Twenty-four patients over a 10-year period were enrolled (mean age, 4.8 years). Respiratory manifestations at initial evaluation included recurrent bronchitis, wheezing, chronic cough, and pneumonia. The duration of the follow-up ranged from 0.5 to 10.5 years. Sputum cultures revealed the presence of Haemophilus influenzae (10 children), Staphylococcus aureus (4 children), and Pseudomonas aeruginosa (3 children). Pancreatic insufficiency was found in two patients. Analysis of the entire coding sequence allowed identification of 16 known mutations in CFTR gene. Fifteen chromosomes (31.2%) carried a mutation in CFTR gene and one allele carried two mutations. Three patients were homozygous or double heterozygous (DeltaF508/DeltaF508, DeltaF508/3849 + 10 kb C-->T, S1235R/G551D). The 5-thymidine allele was identified in four children. CONCLUSION: These results indicate an higher frequency of CFTR gene mutations in patients with borderline sweat chloride test results, compared to data reported in the general population. They lead to the recommendations for complete pulmonary and GI investigations in this group of patients, as well as assiduous care and medical follow-up.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
92 Genotype Poly T 1 -/- 7T/7T 2 R117C/- 7T/7T 3 R75X-D1270H/- 7T/7T 4 -/- 7T/7T 5 G91R/- 7T/5T 6 ⌬F508/- 7T/9T 7 -/- 7T/7T 8 -/- 7T/7T 9 S1235R/G551D 5T/7T 10 ⌬F508/- 9T/9T 11 7T/7T 12 ⌬F508/⌬F508 9T/9T 13 ⌬F508/- 7T/9T 14 -/- 7T/7T 15 ⌬F508/- 7T/9T 16 -/- 7T/5T 17 -/- 7T/7T 18 -/- 7T/7T 19 -/- 7T/9T 20 ⌬F508/- 7T/9T 21 -/- 7T/7T 22 W1282X/- 7T/5T 23 -/- 7T/7T 24 ⌬F508/3849 ϩ 10 kb C 3 T 7T/7T 1594 Clinical Investigations reported in the general population (frequency of the 5T allele in the general population, 5.2%).26 Based on the results of DNA analysis and according to the consensus statement on the diagnosis of CF, three patients (patients 9, 12, and 24) met the criteria of both respiratory manifestations and identification of two CF mutations.21 For patient 6, there was a diagnostic dilemma. Login to comment

[hide] Improved detection of cystic fibrosis mutations in... Genet Med. 2001 May-Jun;3(3):168-76. Heim RA, Sugarman EA, Allitto BA
Improved detection of cystic fibrosis mutations in the heterogeneous U.S. population using an expanded, pan-ethnic mutation panel.
Genet Med. 2001 May-Jun;3(3):168-76., [PMID:11388756]

Abstract [show]
PURPOSE: To determine the comparative frequency of 93 CFTR mutations in U.S. individuals with a clinical diagnosis of cystic fibrosis (CF). METHODS: A total of 5,840 CF chromosomes from Caucasians, Ashkenazi Jews, Hispanics, African Americans, Native Americans, Asians, and individuals of mixed race were analyzed using a pooled ASO hybridization strategy. RESULTS: Sixty-four mutations provided a sensitivity of 70% to 95% in all ethnic groups except Asians, and at least 81% when the U.S. population was considered as a whole. CONCLUSIONS: For population-based carrier screening for CF in the heterogeneous U.S. population, which is characterized by increasing admixture, a pan-ethnic mutation panel of 50 to 70 CFTR mutations may provide a practical test that maximizes sensitivity.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
87 These were G91R, 711 ϩ 5GϾA, T338I, 712-1GϾT, Q359K/T360K, 1161delC, 1609delCA, S549I, Q552X, 1949del84, 1989 ϩ 5GϾT, S1251N, and R1283M. Login to comment

[hide] Prenatal detection of cystic fibrosis by ultrasono... J Med Genet. 2002 Jun;39(6):443-8. Scotet V, De Braekeleer M, Audrezet MP, Quere I, Mercier B, Dugueperoux I, Andrieux J, Blayau M, Ferec C
Prenatal detection of cystic fibrosis by ultrasonography: a retrospective study of more than 346 000 pregnancies.
J Med Genet. 2002 Jun;39(6):443-8., [PMID:12070257]

Abstract [show]

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
246 Therefore, the spectrum of mutations identified in this fetal population is not representative of that identified in our CF population, in which we found a significant number of mild mutations or mutations for which the clinical consequences are not yet established (for example, G91R, R117H, R347L, R560K).34 35 These findings provide the foundation for further investigations towards understanding the pathogenesis of early bowel disease, but also why it results in manifestations in the bowel rather than in the lungs during the fetal period. Login to comment

[hide] Genotype-phenotype correlation in cystic fibrosis:... Am J Med Genet. 2002 Jul 22;111(1):88-95. Salvatore F, Scudiero O, Castaldo G
Genotype-phenotype correlation in cystic fibrosis: the role of modifier genes.
Am J Med Genet. 2002 Jul 22;111(1):88-95., 2002-07-22 [PMID:12124743]

Abstract [show]
More than 1,000 mutations have been identified in the cystic fibrosis (CF) transmembrane regulator (CFTR) disease gene. The impact of these mutations on the protein and the wide spectrum of CF phenotypes prompted a series of Genotype-Phenotype correlation studies. The CFTR genotype is invariably correlated with pancreatic status-in about 85% of cases with pancreatic insufficiency and in about 15% of cases with pancreatic sufficiency. The correlations between the CFTR genotype and pulmonary, liver, and gastrointestinal expression are debatable. The heterogeneous phenotype in CF patients bearing the same genotype or homozygotes for nonsense mutations implicated environmental and/or genetic factors in the disease. However, the discordant phenotype observed in CF siblings argued against a major role of environmental factors and suggested that genes other than CFTR modulate the CF phenotype. A locus that modulates gastrointestinal expression was identified in mice and subsequently in humans. By analyzing nine CF patients discordant for meconium ileus we were able to show that this locus had a dominant effect. Moreover, in a collaborative study we found a higher rate of polymorphisms in beta-defensin genes 1 and 2 in CF patients and in controls. In another multicenter study mutations in alpha-1 antitrypsin (A1AT) and mannose binding lectin genes were found to be independent risk factors for liver disease in CF patients. The body of evidence available suggests that the variegated CF phenotype results from complex interactions between numerous gene products.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
46 A series of mutations usually associated with pancreatic sufficiency have been identified and defined as ''mild`` with reference to pancreatic status [Kerem et al., 1989c]: G85E, G91R, R117H, E193K, P205S, R334W, T338I, R347H, R347L, R347P, R352Q, A455E, S492F, S549N, P574H, D579G, 711 þ 5 G > A, C866Y, F1052V, H1054D, R1066H, R1068H, H1085R, D1152H, S1159P, S1251N, F1286S, G1349D, 2789 þ 5 G > A, and 3849 þ 10kb C > T [Dean et al., 1990; Cutting et al., 1990a; Cremonesi et al., 1992; Highsmith et al., 1994]. Login to comment

[hide] Spatial and temporal distribution of cystic fibros... Hum Genet. 2002 Sep;111(3):247-54. Epub 2002 Aug 1. Scotet V, Gillet D, Dugueperoux I, Audrezet MP, Bellis G, Garnier B, Roussey M, Rault G, Parent P, De Braekeleer M, Ferec C
Spatial and temporal distribution of cystic fibrosis and of its mutations in Brittany, France: a retrospective study from 1960.
Hum Genet. 2002 Sep;111(3):247-54. Epub 2002 Aug 1., [PMID:12215837]

Abstract [show]
Cystic fibrosis (CF) is the most common severe inherited disorder that affects children in Caucasian populations. The aim of this study was to define the spatial and temporal distribution of CF and its mutations in Brittany (western France) where the frequency of the disease is high. We retrospectively registered all CF patients born in Brittany since 1960 by cross-checking various data sources (e.g. medical care centres, genetics laboratories, hospital archives). Councils were contacted so that the place of residence of patients at birth could be determined. Moreover, the spectrum of CF transmembrane conductance regulator (CFTR) mutations and their spatial distribution across Brittany were determined. A total of 520 patients was registered in this study. The incidence of CF was assessed according to administrative (department, district) and diocesan divisions of Brittany and its evolution analysed over four decades. The incidence of CF was 1/2630, with a west/east gradient that was confirmed over time (Finistere: 1/2071 vs Ille-et-Vilaine: 1/3286). At present, the incidence of CF is decreasing, mainly as a result of prenatal diagnosis. An excellent mutation detection rate of 99.7% was obtained. Western Brittany presented a specific spectrum of mutations: 1078delT (9.4% of mutated alleles in the diocese of Cornouaille), G551D (7.7% in the diocese of Leon), 4005+1G-->A (2.9% in Cornouaille) and W846X (1.5% in western Brittany). On the other hand, the eastern region showed a spectrum more similar to the overall picture in France as a whole. This study enabled a precise measurement of the incidence of CF in Brittany to be obtained. The high frequency of the CFTR mutated alleles may result from founder effects and genetic drifts. Moreover, the study brings together the regional specificities of the CFTR gene and highlights disparities that exist in this part of France, both in incidence and in mutation distribution. These are attributable to different degrees of isolation and of population movements between the eastern and western parts of the region. Given that this is the first time that such a detailed study of the CFTR gene has been performed on a large population, this heightened knowledge of the epidemiology of CF in Brittany should provide a basis for the improvement of diagnostic strategies and refinement of genetic counselling.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
118 His genotype was ∆F508/∆F508 Mutation Exon Basse-Bretagne Haute-Bretagne Brittanya ∆F508 10 446 75.6% 224 73.7% 672 75.0% 1078delT 7 31 5.3% 3 1.0% 34 3.8% G551D 11 21 3.6% 12 3.9% 33 3.7% N1303K 21 3 0.5% 9 3.0% 12 1.3% W846X 14a 9 1.5% 1 0.3% 10 1.1% 2789+5G→A 14b 3 0.5% 6 2.0% 9 1.0% 1717-1G→A 11 5 0.8% 3 1.0% 8 0.9% Y1092X 17b 1 0.2% 6 2.0% 7 0.8% 4005+1G→A 20 6 1.0% 1 0.3% 7 0.8% E60X 3 3 0.5% 3 1.0% 6 0.7% 621+1G→T 4 3 0.5% 3 1.0% 6 0.7% R347H 7 6 1.0% 0 0.0% 6 0.7% S492F 10 2 0.3% 3 1.0% 5 0.6% G542X 11 4 0.7% 1 0.3% 5 0.6% 3272-26A→G 17b 2 0.3% 3 1.0% 5 0.6% R117H 4 3 0.5% 1 0.3% 4 0.4% G91R 3 3 0.5% 0 0.0% 3 0.3% ∆I507 10 1 0.2% 2 0.7% 3 0.3% R553X 11 3 0.5% 0 0.0% 3 0.3% W1282X 20 2 0.3% 1 0.3% 3 0.3% A72D 3 0 0.0% 2 0.7% 2 0.2% G85E 3 0 0.0% 2 0.7% 2 0.2% F311L 7 0 0.0% 2 0.7% 2 0.2% 1221delCT 7 2 0.3% 0 0.0% 2 0.2% R560K 11 0 0.0% 2 0.7% 2 0.2% 2622+1G→A 13 2 0.3% 0 0.0% 2 0.2% S945L 15 0 0.0% 2 0.7% 2 0.2% I1234V 19 2 0.3% 0 0.0% 2 0.2% G1249R 20 2 0.3% 0 0.0% 2 0.2% 3905insT 20 2 0.3% 0 0.0% 2 0.2% Unidentified - 3 0.5% 0 0.0% 3 0.3% Total - 590 65.7% 304 34.3% 896 100% IVS17bTA, IVS17bCA) of Irish, Scottish, English, Breton and Czech subjects who were carriers of this mutation, and showed that all these alleles carried a unique haplotype (16-7-17), testifying to the Celtic origin of this mutation (Cashman et al. 1995). Login to comment

[hide] Comparison of the CFTR mutation spectrum in three ... Hum Mutat. 2003 Jul;22(1):105. Scotet V, Barton DE, Watson JB, Audrezet MP, McDevitt T, McQuaid S, Shortt C, De Braekeleer M, Ferec C, Le Marechal C
Comparison of the CFTR mutation spectrum in three cohorts of patients of Celtic origin from Brittany (France) and Ireland.
Hum Mutat. 2003 Jul;22(1):105., [PMID:12815607]

Abstract [show]
This study aims to compare the spectrum of the mutations identified in the gene responsible for cystic fibrosis in three cohorts of patients of Celtic origin from Brittany and Ireland. It included 389 patients from Brittany, 631 from Dublin and 139 from Cork. The CFTR gene analysis relied on the detection of the most common mutations, followed by a complete gene scanning using DGGE or D-HPLC. High mutation detection rates were obtained in each cohort: 99.6%, 96.8%, and 96.0% respectively. A high frequency of the c.1652_1655 del3 mutation (F508del: 74.8% to 81.3%) and of the "Celtic" mutation (c.1784G>A (G551D): 3.7% to 9.7%) was observed in each population. Apart from this, the mutation spectrums differed. In Brittany, the most common abnormalities were: c.1078delT (3.6%), c.4041C>G (N1303K: 1.4%), c.2670G>A (W846X(2): 1.0%) and c.1717-1G>A (1.0%), whereas in the cohort of Dublin, the main mutations were: c.482G>A (R117H: 3.0%), c.1811G>C (R560T: 2.4%) and c.621+1G>T (1.7%). Finally, in the Cork area, only the c.482G>A mutation (R117H) reached a frequency of 1%. Two previously-unreported mutations were identified in the Dublin cohort: c.2623-2A>G and c.3446T>G (M1105R). This collaborative study highlights the similarities of the CFTR alleles in the Breton and Irish populations, but also the disparities that exist between these populations, despite their common origin. Each population has its own history, with its mixture of founder effects and genetic drifts, which are at the origin of the current mutation distribution. The molecular study of the CFTR gene provides new tools for retracing European populations' histories.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
64 Spectrum of the CFTR Mutations Identified in the Cohorts from Brittany, Dublin Centre, and Cork Area Nucleotide Amino acid change * change Exon Number Frequency Number Frequency Number Frequency 211delG 2 1 0.1% 310G>T E60X 3 5 0.6% 4 0.3% 347C>A A72D 3 1 0.1% 368G>A W79X 3 1 0.1% 386G>A G85E 3 2 0.3% 3 0.2% 403G>A G91R 3 2 0.3% 482G>A R117H 4 4 0.5% 38 3.0% 4 1.4% 498T>A Y122X 4 1 0.1% 574delA 4 1 0.1% 577G>A G149R 4 1 0.1% 621+1G>T int 4 5 0.6% 21 1.7% 790C>T Q220X 6a 1 0.1% 875+1G>C int 6a 1 0.4% 905delG 6b 1 0.1% 1065C>G F311L 7 2 0.3% 1078delT 7 28 3.6% 1132C>T R334W 7 1 0.1% 1172G>A R347H 7 5 0.6% 1172G>T R347L 7 1 0.1% 1172G>C R347P 7 1 0.1% 1187G>A R352Q 7 3 0.2% 2 0.7% 1208A>G Q359R 7 1 0.1% 1154insTC 7 2 0.2% 1221delCT 7 2 0.3% 1248+1G>A int 7 1 0.1% 1249-27delTA int 7 1 0.4% 1334G>A W401X 8 1 0.1% 1461ins4 9 5 0.4% 1471delA 9 2 0.2% 1607C>T S492F 10 2 0.3% 1609C>T Q493X 10 1 0.1% 1648_1653delATC I507del 10 3 0.4% 10 0.8% 1 0.4% 1652_1655del 3 bp F508del 10 582 74.8% 966 76.5% 226 81.3% 1690G>T V520F 10 4 0.3% 1717-1G>A int 10 8 1.0% 9 0.7% 1756G>T G542X 11 5 0.6% 8 0.6% 1779T>G S549R 11 1 0.1% 1784G>A G551D 11 29 3.7% 82 6.5% 27 9.7% 1789C>G R553G 11 1 0.1% 1789C>T R553X 11 3 0.4% 1 0.1% 1806delA 11 1 0.1% 1811G>A R560K 11 2 0.3% 1811G>C R560T 11 30 2.4% 2 0.7% 1819T>A Y563N 12 1 0.1% 1853C>A P574H 12 1 0.1% 1898+1G>A int 12 1 0.1% 2184delA 13 1 0.1% 1 0.1% 2184insA 13 1 0.1% 2622+1G>A int 13 1 0.1% 2 0.2% 2622+1G>T int 13 1 0.1% 2623-2A>G ** int 13 1 0.1% 2670G>A W846X2 14a 8 1.0% 2752-1G>T int 14a 1 0.1% 2752-26A>G int 14a 2 0.2% 2789+5G>A int 14b 6 0.8% 2966C>T S945L 15 2 0.3% 3007delG 15 4 0.3% 3040G>C G970R 15 1 0.1% 3062C>T S977F 16 1 0.1% 3120+1G>A int 16 1 0.1% 3272-26A>G int 17a 4 0.5% 2 0.2% 2 0.7% 3320dupli(CTATG) 17b 1 0.1% 3329G>A R1066H 17b 1 0.1% 3340C>T R1070W 17b 1 0.1% 3408C>A Y1092X 17b 7 0.9% 3442G>T E1104X 17b 1 0.1% 3446T>G ** M1105R 17b 1 0.1% 3586G>C D1152H 18 1 0.1% 3601-17T>C + 1367delC int 18 + 9 1 0.1% 3616C>T R1162X 19 1 0.1% 2 0.2% 3659delC 19 2 0.2% 3832A>G I1234V 19 2 0.3% 3849+4A>G int 19 1 0.1% 3849+10kbC>T int 19 3 0.2% 3877G>A G1249R 20 1 0.1% 3884G>A S1251N 20 1 0.1% 3898insC 20 1 0.1% 3905insT 20 2 0.3% 3978G>A W1282X 20 3 0.4% 4005+1G>A int 20 6 0.8% 4016insT 21 1 0.1% 4041C>G N1303K 21 11 1.4% 5 0.4% 4136T>C L1335P 22 1 0.1% 1 0.4% 4279insA 23 1 0.1% Unidentified Unidentified - 3 0.4% 41 3.2% 11 4.0% Total 778 100.0% 1262 100.0% 278 100.0% * All nucleotide changes correspond to cDNA numbering. Login to comment

[hide] CFTR genotypes in patients with normal or borderli... Hum Mutat. 2003 Oct;22(4):340. Feldmann D, Couderc R, Audrezet MP, Ferec C, Bienvenu T, Desgeorges M, Claustres M, Mittre H, Blayau M, Bozon D, Malinge MC, Monnier N, Bonnefont JP, Iron A, Bieth E, Dumur V, Clavel C, Cazeneuve C, Girodon E
CFTR genotypes in patients with normal or borderline sweat chloride levels.
Hum Mutat. 2003 Oct;22(4):340., [PMID:12955726]

Abstract [show]
In recent years, some patients bearing "atypical" forms of cystic fibrosis (CF) with normal sweat chloride concentrations have been described. To identify the spectrum of mutant combinations causing such atypical CF, we collected the results of CFTR (ABCC7) mutation analysis from 15 laboratories. Thirty patients with one or more typical symptoms of the disease associated with normal or borderline sweat chloride levels and bearing two CFTR mutations were selected. Phenotypes and genotypes of these 30 patients are described. A total of 18 different CFTR mutations were observed in the 60 chromosomes analysed. F508del was present in 31.6 % of the mutated chromosomes and 3849+10kbC>T in 13.3 %. R117H, D1152H, L206W, 3272-26A>G, S1235R, G149R, R1070W, S945L, and the poly-T tract variation commonly called IVS8-5T were also observed. The relative frequency of CFTR mutations clearly differed from that observed in typical CF patients or in CBAVD patients with the same ethnic origin. A mild genotype with one or two mild or variable mutations was observed in all the patients. These findings improve our understanding of the distribution of CFTR alleles in CF with normal or borderline sweat chloride concentrations and will facilitate the development of more sensitive CFTR mutation screening.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
44 Table 1 : Genotypes and Phenotypes of Patients with Normal or BordIerline Sweat Tests Patient Age at diagnosis (years) CFTR GENOTYPE* Allele 1 Allele 2 SWEAT CL- MEAN (MMOL/L) PHENOTYPE 1 0.2 F508del G149R 38 P+PI, neonatal hypertrypsinemia, 2 0.3 G551D R117H-7T 31 neonatal hypertrypsinemia 3 0.4 F508del R1070W 30.5 neonatal hypertrypsinemia 4 0.4 F508del R117H-7T 52 P 5 0.6 F508del 3849+10kbC>T 48 P 6 0.11 F508del S945L 58 P+PI 7 1 F508del 5T 40 P+CBAVD 8 2 F508del L206W 53 P 9 2 W1282X 5T 42.5 P 10 5 F508del 3849+10kbC>T 55.5 P 11 5 F508del L206W 55 P 12 5 G91R 5T 47.5 P 13 6 G551D S1235R+5T 49.5 P, neonatal hypertrypsinemia 14 7 F508del 3849+10kb 50 P, nasal popyposis 15 13 F508del R117H-7T 58 P, nasal polyposis 16 18 F508del 5T 60.5 P 17 20 G542X 3849+10kbC>T 52 P+PI 18 21 I507del 3849+10kbC>T 54 P, bronchiectasis 19 30 R347P 3849+10kbC>T 43 P, Pseudomonas colonisation 20 30 I507del L206W 57.5 CBAVD, chronic cough 21 31 F508del R117H-7T 60 CBAVD 22 32 G542X 3849+10kbC>T 30 P, Pseudomonas colonisation 23 34 F508del 3272-26A>G 64 P, CBAVD 24 37 R1070Q D1152H 56 CBAVD, bronchectasis 25 46 F508del D1152H 43 P 26 55 F508del D1152H 48 P, Pseudomonas colonisation 27 56 I507del S1235R 53 P 28 >18 F508del D1152H 60 P+PI 29 >20 F508del 3849+10kbC>T 18 P, bronchiectasis 30 >20 F508del 3272-26A>G 61 P *All mutations are named in accordance with the numbering used in the CFTR Mutation Database: http://www.genet.sickkids.on.ca/cftr/. Login to comment

[hide] Relation of sweat chloride concentration to severi... Pediatr Pulmonol. 2004 Sep;38(3):204-9. Davis PB, Schluchter MD, Konstan MW
Relation of sweat chloride concentration to severity of lung disease in cystic fibrosis.
Pediatr Pulmonol. 2004 Sep;38(3):204-9., [PMID:15274098]

Abstract [show]
In cystic fibrosis (CF), sweat chloride concentration has been proposed as an index of CFTR function for testing systemic drugs designed to activate mutant CFTR. This suggestion arises from the assumption that greater residual CFTR function should lead to a lower sweat chloride concentration, as well as protection against severe lung disease. This logic gives rise to the hypothesis that the lower the sweat chloride concentration, the less severe the lung disease. In order to test this hypothesis, we studied 230 patients homozygous for the DeltaF508 allele, and 34 patients with at least one allele associated with pancreatic sufficiency, born since January 1, 1955, who have pulmonary function data and sweat chloride concentrations recorded in our CF center database, and no culture positive for B. cepacia. We calculated a severity index for pulmonary disease, using an approach which takes into account all available pulmonary function data as well as the patient's current age and survival status. Patients with alleles associated with pancreatic sufficiency had significantly better survival (P = 0.0083), lower sweat chloride concentration (81.4 +/- 23.8 vs. 103.2 +/- 14.2 mEq/l, P < 0.0001), slower rate of decline of FEV(1) % predicted (-0.75 +/- 0.34 vs. -2.34 +/- 0.17% predicted per year), and a better severity index than patients homozygous for the DeltaF508 allele (median 73rd percentile vs. median 55th percentile, P = 0.0004). However, the sweat chloride concentration did not correlate with the severity index, either in the population as a whole, or in the population of patients with alleles associated with pancreatic sufficiency, who are thought to have some residual CFTR function. These data suggest that, by itself, sweat chloride concentration does not necessarily predict a milder pulmonary course in patients with cystic fibrosis.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
27 T; G91R; E92K; P205S; G551S; Y563N; and P574H.23,24 Note that there are 36 mild alleles in 34 subjects, because two subjects had both the 3848 þ 10 kb C ! Login to comment

[hide] CFTR mutation distribution among U.S. Hispanic and... Genet Med. 2004 Sep-Oct;6(5):392-9. Sugarman EA, Rohlfs EM, Silverman LM, Allitto BA
CFTR mutation distribution among U.S. Hispanic and African American individuals: evaluation in cystic fibrosis patient and carrier screening populations.
Genet Med. 2004 Sep-Oct;6(5):392-9., [PMID:15371903]

Abstract [show]
PURPOSE: We reviewed CFTR mutation distribution among Hispanic and African American individuals referred for CF carrier screening and compared mutation frequencies to those derived from CF patient samples. METHODS: Results from CFTR mutation analyses received from January 2001 through September 2003, were analyzed for four populations: Hispanic individuals with a CF diagnosis (n = 159) or carrier screening indication (n = 15,333) and African American individuals with a CF diagnosis (n = 108) or carrier screening indication (n = 8,973). All samples were tested for the same 87 mutation panel. RESULTS: In the Hispanic population, 42 mutations were identified: 30 in the patient population (77.5% detection rate) and 33 among carrier screening referrals. Five mutations not included in the ACMG/ACOG carrier screening panel (3876delA, W1089X, R1066C, S549N, 1949del84) accounted for 7.55% detection in patients and 5.58% among carriers. Among African American referrals, 33 different mutations were identified: 21 in the patient population (74.4% detection) and 23 in the carrier screening population. Together, A559T and 711+5G>A were observed at a detection rate of 3.71% in CF patients and 6.38% in carriers. The mutation distribution seen in both the carrier screening populations reflected an increased frequency of mutations with variable expression such as D1152H, R117H, and L206W. CONCLUSIONS: A detailed analysis of CFTR mutation distribution in the Hispanic and African American patient and carrier screening populations demonstrates that a diverse group of mutations is most appropriate for diagnostic and carrier screening in these populations. To best serve the increasingly diverse U.S. population, ethnic-specific mutations should be included in mutation panels.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
35 87 mutation panel The following mutations were included in the panel: ⌬F508, ⌬F311, ⌬I507, A455E, A559T, C524X, D1152H, D1270N, E60X, G178R, G330X, G480C, G542X, G551D, G85E, G91R, I148T, K710X, L206W, M1101K, N1303K, P574H, Q1238X, Q359K/T360K, Q493X, Q552X, Q890X, R1066C, R1158X, R1162X, R117C, R117H, R1283M, R334W, R347H, R347P, R352Q, R553X, R560T, S1196X, S1251N, S1255X, S364P, S549I, S549N, S549R, T338I, V520F, W1089X, W1282X, Y1092X, Y563D, 1078delT, 1161delC, 1609delCA, 1677delTA, 1717-1GϾA, 1812-1GϾA, 1898ϩ1GϾA, 1898ϩ5GϾT, 1949del84, 2043delG, 2143delT, 2183delAAϾG, 2184delA, 2307insA, 2789ϩ5GϾA, 2869insG, 3120ϩ1GϾA, 3120GϾA, 3659delC, 3662delA, 3791delC, 3821delT, 3849ϩ10kbCϾT, 3849ϩ4AϾG, 3905insT, 394delTT, 405ϩ1GϾA, 405ϩ3AϾC, 444delA, 574delA, 621ϩ1GϾT, 711ϩ1GϾT, 711ϩ5GϾA, 712-1GϾT, 3876delA CFTR mutation analysis Genomic DNA was extracted from peripheral blood lymphocytes, buccal cell swabs, or bloodspots by Qiagen QIAmp 96 DNA Blood Kit. Specimens were tested for 87 mutations by a pooled allele-specific oligonucleotide (ASO) hybridization method as previously described.16,17 Two multiplex chain reactions (PCR) were used to amplify 19 regions of the CFTR gene. Login to comment

[hide] A foldable CFTR{Delta}F508 biogenic intermediate a... J Cell Biol. 2004 Dec 20;167(6):1075-85. Younger JM, Ren HY, Chen L, Fan CY, Fields A, Patterson C, Cyr DM
A foldable CFTR{Delta}F508 biogenic intermediate accumulates upon inhibition of the Hsc70-CHIP E3 ubiquitin ligase.
J Cell Biol. 2004 Dec 20;167(6):1075-85., 2004-12-20 [PMID:15611333]

Abstract [show]
CFTRDeltaF508 exhibits a correctable protein-folding defect that leads to its misfolding and premature degradation, which is the cause of cystic fibrosis (CF). Herein we report on the characterization of the CFTRDeltaF508 biogenic intermediate that is selected for proteasomal degradation and identification of cellular components that polyubiquitinate CFTRDeltaF508. Nonubiquitinated CFTRDeltaF508 accumulates in a kinetically trapped, but folding competent conformation, that is maintained in a soluble state by cytosolic Hsc70. Ubiquitination of Hsc70-bound CFTRDeltaF508 requires CHIP, a U box containing cytosolic cochaperone. CHIP is demonstrated to function as a scaffold that nucleates the formation of a multisubunit E3 ubiquitin ligase whose reconstituted activity toward CFTR is dependent upon Hdj2, Hsc70, and the E2 UbcH5a. Inactivation of the Hsc70-CHIP E3 leads CFTRDeltaF508 to accumulate in a nonaggregated state, which upon lowering of cell growth temperatures, can fold and reach the cell surface. Inhibition of CFTRDeltaF508 ubiquitination can increase its cell surface expression and may provide an approach to treat CF.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
451 Structural cues involved in endoplasmic reticulum degradation of G85E and G91R mutant cystic fibrosis transmembrane conductance regulator. J. Clin. Login to comment

[hide] Side chain and backbone contributions of Phe508 to... Nat Struct Mol Biol. 2005 Jan;12(1):10-6. Epub 2004 Dec 26. Thibodeau PH, Brautigam CA, Machius M, Thomas PJ
Side chain and backbone contributions of Phe508 to CFTR folding.
Nat Struct Mol Biol. 2005 Jan;12(1):10-6. Epub 2004 Dec 26., [PMID:15619636]

Abstract [show]
Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR), an integral membrane protein, cause cystic fibrosis (CF). The most common CF-causing mutant, deletion of Phe508, fails to properly fold. To elucidate the role Phe508 plays in the folding of CFTR, missense mutations at this position were generated. Only one missense mutation had a pronounced effect on the stability and folding of the isolated domain in vitro. In contrast, many substitutions, including those of charged and bulky residues, disrupted folding of full-length CFTR in cells. Structures of two mutant nucleotide-binding domains (NBDs) reveal only local alterations of the surface near position 508. These results suggest that the peptide backbone plays a role in the proper folding of the domain, whereas the side chain plays a role in defining a surface of NBD1 that potentially interacts with other domains during the maturation of intact CFTR.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
337 Xiong, X., Bragin, A., Widdicombe, J.H., Cohn, J. & Skach, W.R. Structural cues involved in endoplasmic reticulum degradation of G85E and G91R mutant cystic fibrosis transmembrane conductance regulator. J. Clin. Login to comment
336 Xiong, X., Bragin, A., Widdicombe, J.H., Cohn, J. & Skach, W.R. Structural cues involved in endoplasmic reticulum degradation of G85E and G91R mutant cystic fibrosis transmembrane conductance regulator. J. Clin. Login to comment

[hide] Pharmacological induction of CFTR function in pati... Pediatr Pulmonol. 2005 Sep;40(3):183-96. Kerem E
Pharmacological induction of CFTR function in patients with cystic fibrosis: mutation-specific therapy.
Pediatr Pulmonol. 2005 Sep;40(3):183-96., [PMID:15880796]

Abstract [show]
CFTR mutations cause defects of CFTR protein production and function by different molecular mechanisms. Mutations can be classified according to the mechanisms by which they disrupt CFTR function. This understanding of the different molecular mechanisms of CFTR dysfunction provides the scientific basis for the development of targeted drugs for mutation-specific therapy of cystic fibrosis (CF). Class I mutations are nonsense mutations that result in the presence of a premature stop codon that leads to the production of unstable mRNA, or the release from the ribosome of a short, truncated protein that is not functional. Aminoglycoside antibiotics can suppress premature termination codons by disrupting translational fidelity and allowing the incorporation of an amino acid, thus permitting translation to continue to the normal termination of the transcript. Class II mutations cause impairment of CFTR processing and folding in the Golgi. As a result, the mutant CFTR is retained in the endoplasmic reticulum (ER) and eventually targeted for degradation by the quality control mechanisms. Chemical and molecular chaperones such as sodium-4-phenylbutyrate can stabilize protein structure, and allow it to escape from degradation in the ER and be transported to the cell membrane. Class III mutations disrupt the function of the regulatory domain. CFTR is resistant to phosphorylation or adenosine tri-phosphate (ATP) binding. CFTR activators such as alkylxanthines (CPX) and the flavonoid genistein can overcome affected ATP binding through direct binding to a nucleotide binding fold. In patients carrying class IV mutations, phosphorylation of CFTR results in reduced chloride transport. Increases in the overall cell surface content of these mutants might overcome the relative reduction in conductance. Alternatively, restoring native chloride pore characteristics pharmacologically might be effective. Activators of CFTR at the plasma membrane may function by promoting CFTR phosphorylation, by blocking CFTR dephosphorylation, by interacting directly with CFTR, and/or by modulation of CFTR protein-protein interactions. Class V mutations affect the splicing machinery and generate both aberrantly and correctly spliced transcripts, the levels of which vary among different patients and among different organs of the same patient. Splicing factors that promote exon inclusion or factors that promote exon skipping can promote increases of correctly spliced transcripts, depending on the molecular defect. Inconsistent results were reported regarding the required level of corrected or mutated CFTR that had to be reached in order to achieve normal function.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
58 C-D565G II DF508 D1507 S549R S549I S549N S549R S945D S945L H1054D G1061R L1065P R1066C R1066M L1077P H1085R N1303K G85E III G551D S492F V520F R553G R560T R560S Y569D IV R117H, R117C, R117P, R117L D1152H, L88S, G91R, E92K, Q98R, P205S, L206W, L227R, F311L, G314E, R334W, R334Q, I336K, T338I, L346P, R347C, R347H, R347L, R347P, L927P, R1070W, R1070Q V 3849 þ 10 kb C ! Login to comment

[hide] Cystic fibrosis. N Engl J Med. 2005 May 12;352(19):1992-2001. Rowe SM, Miller S, Sorscher EJ
Cystic fibrosis.
N Engl J Med. 2005 May 12;352(19):1992-2001., 2005-05-12 [PMID:15888700]

Abstract [show]

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
119 Like ∆F508, several other clinically important mutations - such as N1303K, G85E, and G91R - lead to misfolded CFTR protein that is prematurely degraded. Login to comment

[hide] Genetics of cystic fibrosis. Semin Respir Crit Care Med. 2003 Dec;24(6):629-38. Gallati S
Genetics of cystic fibrosis.
Semin Respir Crit Care Med. 2003 Dec;24(6):629-38., [PMID:16088579]

Abstract [show]
Cystic fibrosis (CF) is caused by mutations in the CF transmembrane conductance regulator (CFTR) gene, which encodes a protein expressed in the apical membrane of exocrine epithelial cells. CFTR functions principally as a cyclic adenosine monophosphate (cAMP)-induced chloride channel and appears capable of regulating other ion channels. Mutations affect CFTR through a variety of molecular mechanisms, which can produce little or no functional gene product at the apical membrane. More than 1000 different disease-causing mutations within the CFTR gene have been described. The potential of a mutation to contribute to the phenotype depends on its type, localization in the gene, and the molecular mechanism as well as on interactions with secondary modifying factors. Genetic testing can confirm a clinical diagnosis of CF and can be used for infants with meconium ileus, for carrier detection in individuals with positive family history and partners of proven CF carriers, and for prenatal diagnostic testing if both parents are carriers. Studies of clinical phenotype in correlation with CFTR genotype have revealed a very complex relationship demonstrating that some phenotypic features are closely determined by the underlying mutations, whereas others are modulated by modifier genes, epigenetic mechanisms, and environment.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
50 In effect, virtually no func- Table 2 Unusually Common Cystic Fibrosis Mutations in Specific Populationsa Total Exon/ Number Number Frequency Mutation Intron Ethnic Origin Observed Screened (%) 296+12T→C intron 02 Pakistani 02 24 8.33 E60X exon 03 Belgian 06 394 1.52 G91R exon 03 French 04 266 1.50 394delTT exon 03 Scandinavian 78 1588 4.91 457TAT→G exon 04 Austrian 04 334 1.20 Y122X exon 04 Réunion Island 14 29 48.27 I148T exon 04 French Canadian 06 66 9.09 711+5G→A intron 05 Italian (North East) 06 225 2.67 1078delT exon 07 Celtic 27 475 5.68 1161delC exon 07 Pakistani 02 24 8.33 T338I exon 07 Italian, Sardinian 04 86 4.65 Q359K/T360K exon 07 Georgian Jews 07 8 87.50 R347H exon 07 Turkish 04 134 2.98 1609delCA exon 10 Spanish 03 96 3.12 1677delTA exon 10 Bulgarian 05 222 2.25 S549I exon 11 Arabs 02 40 5.00 Q552X exon 11 Italian (North East) 03 225 1.33 A559T exon 11 African-American 02 79 2.53 1811+1.2kbA→G intron 11 Spanish 22 1068 2.06 1898+5G→T intron 12 Chinese 03 10 30.00 1949del84 exon 13 Spanish 02 136 1.47 2143delT exon 13 Russian 04 118 3.39 2183AA→G exon 13 Italian (North East) 21 225 9.33 2184insA exon 13 Russian 03 118 2.54 3120+1G→A intron 16 African-American 14 112 12.50 3272-26A→G intron 17a Portugese, French 06 386 1.55 R1066C exon 17b Portugese 05 105 4.76 R1070Q exon 17b Bulgarian 04 166 2.41 Y1092X exon 17b French Canadian, 11 725 1.52 French M1101K exon 17b Hutterite 22 32 68.75 3821delT exon 19 Russian 03 118 2.54 S1235R exon 19 French (South) 04 340 1.18 S1251N exon 20 Dutch, Belgian 11 792 1.39 S1255X exon 20 African-American 02 79 2.53 3905insT exon 20 Swiss 45 982 4.58 Amish, Arcadian 13 86 15.12 W1282X Exon 20 Jewish-Ashkenazi 50 95 52.63 R1283M exon 20 Welsh 03 183 1.64 aAccording to the Cystic Fibrosis Genetic Analysis Consortium, http://www.genet.sickkids.on.ca/cftr/. Login to comment

[hide] The role of the UPS in cystic fibrosis. BMC Biochem. 2007 Nov 22;8 Suppl 1:S11. Turnbull EL, Rosser MF, Cyr DM
The role of the UPS in cystic fibrosis.
BMC Biochem. 2007 Nov 22;8 Suppl 1:S11., [PMID:18047735]

Abstract [show]
CF is an inherited autosomal recessive disease whose lethality arises from malfunction of CFTR, a single chloride (Cl-) ion channel protein. CF patients harbor mutations in the CFTR gene that lead to misfolding of the resulting CFTR protein, rendering it inactive and mislocalized. Hundreds of CF-related mutations have been identified, many of which abrogate CFTR folding in the endoplasmic reticulum (ER). More than 70% of patients harbor the DeltaF508 CFTR mutation that causes misfolding of the CFTR proteins. Consequently, mutant CFTR is unable to reach the apical plasma membrane of epithelial cells that line the lungs and gut, and is instead targeted for degradation by the UPS. Proteins located in both the cytoplasm and ER membrane are believed to identify misfolded CFTR for UPS-mediated degradation. The aberrantly folded CFTR protein then undergoes polyubiquitylation, carried out by an E1-E2-E3 ubiquitin ligase system, leading to degradation by the 26S proteasome. This ubiquitin-dependent loss of misfolded CFTR protein can be inhibited by the application of 'corrector' drugs that aid CFTR folding, shielding it from the UPS machinery. Corrector molecules elevate cellular CFTR protein levels by protecting the protein from degradation and aiding folding, promoting its maturation and localization to the apical plasma membrane. Combinatory application of corrector drugs with activator molecules that enhance CFTR Cl- ion channel activity offers significant potential for treatment of CF patients. Publication history: Republished from Current BioData's Targeted Proteins database (TPdb; http://www.targetedproteinsdb.com).

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
34 Other identified class II disease-causing mutations in CFTR include N1303K, G85E and G91R [18]. Login to comment
36 The exact mechanism by which these mutations disrupt folding is not completely clear [21], but both the G85E and G91R mutations have been shown to affect folding due to the insertion of a charged residue in the plane of the lipid bilayer [9]. Login to comment

[hide] Assembly and misassembly of cystic fibrosis transm... Mol Biol Cell. 2008 Nov;19(11):4570-9. Epub 2008 Aug 20. Rosser MF, Grove DE, Chen L, Cyr DM
Assembly and misassembly of cystic fibrosis transmembrane conductance regulator: folding defects caused by deletion of F508 occur before and after the calnexin-dependent association of membrane spanning domain (MSD) 1 and MSD2.
Mol Biol Cell. 2008 Nov;19(11):4570-9. Epub 2008 Aug 20., [PMID:18716059]

Abstract [show]
Cystic fibrosis transmembrane conductance regulator (CFTR) is a polytopic membrane protein that functions as a Cl(-) channel and consists of two membrane spanning domains (MSDs), two cytosolic nucleotide binding domains (NBDs), and a cytosolic regulatory domain. Cytosolic 70-kDa heat shock protein (Hsp70), and endoplasmic reticulum-localized calnexin are chaperones that facilitate CFTR biogenesis. Hsp70 functions in both the cotranslational folding and posttranslational degradation of CFTR. Yet, the mechanism for calnexin action in folding and quality control of CFTR is not clear. Investigation of this question revealed that calnexin is not essential for CFTR or CFTRDeltaF508 degradation. We identified a dependence on calnexin for proper assembly of CFTR's membrane spanning domains. Interestingly, efficient folding of NBD2 was also found to be dependent upon calnexin binding to CFTR. Furthermore, we identified folding defects caused by deletion of F508 that occurred before and after the calnexin-dependent association of MSD1 and MSD2. Early folding defects are evident upon translation of the NBD1 and R-domain and are sensed by the RMA-1 ubiquitin ligase complex.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
239 Yet, the fact that the G91R mutation, and to a lesser extent, inhibition of calnexin function also hinder NBD2 folding suggests that general disruption of MSD assembly prevents proper folding of NBD2. Login to comment

[hide] Cooperative assembly and misfolding of CFTR domain... Mol Biol Cell. 2009 Apr;20(7):1903-15. Epub 2009 Jan 28. Du K, Lukacs GL
Cooperative assembly and misfolding of CFTR domains in vivo.
Mol Biol Cell. 2009 Apr;20(7):1903-15. Epub 2009 Jan 28., [PMID:19176754]

Abstract [show]
The cystic fibrosis transmembrane conductance regulator (CFTR) architecture consists of two membrane spanning domains (MSD1 and -2), two nucleotide binding domains (NBD1 and -2), and a regulatory (R) domain. Several point mutations lead to the channel misprocessing, with limited structural perturbation of the mutant domain. To gain more insight into the basis of CFTR folding defect, the contribution of domain-wise and cooperative domain folding was assessed by determining 1) the minimal domain combination that is recognized as native and can efficiently escape the endoplasmic reticulum (ER) retention and 2) the impact of mutation on the conformational coupling among domains. One-, two-, three-, and most of the four-domain assemblies were retained at the ER. Solubilization mutations, however, rescued the NBD1 processing defect conceivably by thermodynamic stabilization. The smallest folding unit that traversed the secretory pathway was composed of MSD1-NBD1-R-MSD2 as a linear or split polypeptide. Cystic fibrosis-causing missense mutations in the MSD1, NBD1, MSD2, and NBD2 caused conformational defect in multiple domains. We propose that cooperative posttranslational folding is required for domain stabilization and provides a plausible explanation for the global misfolding caused by point mutations dispersed along the full-length CFTR.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
37 MATERIALS AND METHODS Cell Lines Baby hamster kidney (BHK) cells, stably expressing the wt and mutant (G91R, L346P, L1093P, N1303K, ⌬F508, 4D, 1218X, 1158X, and 823X) CFTR with three tandem hemagglutinin (HA)-epitope (3HA) inserted into the fourth extracellular loop, were isolated and maintained as described previously (Sharma et al., 2004; Du et al., 2005). Login to comment
252 Four CF mutations, G91R (Xiong et al., 1997), L346P (Choi et al., 2005), L1093P (Seibert et al., 1996), and N1303K (Gregory et al., 1991), localized to the transmembrane (TM) 1 and TM6 in MSD1, the cytosolic loop (CL) 4, and in the NBD2, respectively (Figure 7a). Login to comment
274 Remarkably, the G91R, L346P, ⌬F508, 4D, and L1093P mutations, regardless of their location, profoundly augmented the protease susceptibility of the NBD2 (ϳ30 kDa), probed with the M3A7 Ab (Figure 7g and Supplemental Figure S7c). Login to comment
301 One of the most important observations of this study is that the NBD2 conformational stability was dramatically impaired regardless the localization of mutations (G91R, L346P, ⌬F508, 4D, and L1093P) in the NBD1, TM1, TM6, or CL4 (Figure 6e). Login to comment

[hide] Mechanisms for rescue of correctable folding defec... Mol Biol Cell. 2009 Sep;20(18):4059-69. Epub 2009 Jul 22. Grove DE, Rosser MF, Ren HY, Naren AP, Cyr DM
Mechanisms for rescue of correctable folding defects in CFTRDelta F508.
Mol Biol Cell. 2009 Sep;20(18):4059-69. Epub 2009 Jul 22., [PMID:19625452]

Abstract [show]
Premature degradation of CFTRDeltaF508 causes cystic fibrosis (CF). CFTRDeltaF508 folding defects are conditional and folding correctors are being developed as CF therapeutics. How the cellular environment impacts CFTRDeltaF508 folding efficiency and the identity of CFTRDeltaF508's correctable folding defects is unclear. We report that inactivation of the RMA1 or CHIP ubiquitin ligase permits a pool of CFTRDeltaF508 to escape the endoplasmic reticulum. Combined RMA1 or CHIP inactivation and Corr-4a treatment enhanced CFTRDeltaF508 folding to 3-7-fold greater levels than those elicited by Corr-4a. Some, but not all, folding defects in CFTRDeltaF508 are correctable. CHIP and RMA1 recognize different regions of CFTR and a large pool of nascent CFTRDeltaF508 is ubiquitinated by RMA1 before Corr-4a action. RMA1 recognizes defects in CFTRDeltaF508 related to misassembly of a complex that contains MSD1, NBD1, and the R-domain. Corr-4a acts on CFTRDeltaF508 after MSD2 synthesis and was ineffective at rescue of DeltaF508 dependent folding defects in amino-terminal regions. In contrast, misfolding caused by the rare CF-causing mutation V232D in MSD1 was highly correctable by Corr-4a. Overall, correction of folding defects recognized by RMA1 and/or global modulation of ER quality control has the potential to increase CFTRDeltaF508 folding and provide a therapeutic approach for CF.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
183 Thus, we asked to what extent can disease related folding defects caused by mutations in MSD1 (G85E, G91R, and V232D), MSD2 (M1137R), and NBD2 (N1303K) be corrected relative to those caused by deletion of F508 in NBD1 (Figure 4A). Login to comment
184 The G85E, G91R, M1137R, and N1303K mutations all hinder folding of the nascent B-form of CFTR via a mechanism that involves insertion of a charged amino acid into an inappropriate region (Gregory et al., 1991; Xiong et al., 1997; Vankeerberghen et al., 1998). Login to comment
189 The CFTR G85E and G91R point mutations are contained within TM1, whereas the V232D mutation lies within TM4 of CFTR`s MSD1 domain. Login to comment
192 Furthermore, chemical treatment of CFTR G91R or CFTR M1137R did not significantly affect the accumulation of the immature B-form; however, the mature C-forms of both CFTR G91R and CFTR M1137R were apparent. Login to comment
316 Corr-4a-dependent rescue of MSD biogenic mutants G91R and M1137R was no greater than that observed with corrector-treated CFTR⌬F508. Login to comment

[hide] Ubiquitin-mediated proteasomal degradation of ABC ... J Pharm Sci. 2011 Sep;100(9):3602-19. doi: 10.1002/jps.22615. Epub 2011 May 12. Nakagawa H, Toyoda Y, Wakabayashi-Nakao K, Tamaki H, Osumi M, Ishikawa T
Ubiquitin-mediated proteasomal degradation of ABC transporters: a new aspect of genetic polymorphisms and clinical impacts.
J Pharm Sci. 2011 Sep;100(9):3602-19. doi: 10.1002/jps.22615. Epub 2011 May 12., [PMID:21567408]

Abstract [show]
The interindividual variation in the rate of drug metabolism and disposition has been known for many years. Pharmacogenomics dealing with heredity and response to drugs is a part of science that attempts to explain variability of drug responses and to search for the genetic basis of such variations or differences. Genetic polymorphisms of drug metabolizing enzymes and drug transporters have been found to play a significant role in the patients' responses to medication. Accumulating evidence demonstrates that certain nonsynonymous polymorphisms have great impacts on the protein stability and degradation, as well as the function of drug metabolizing enzymes and transporters. The aim of this review article is to address a new aspect of protein quality control in the endoplasmic reticulum and to present examples regarding the impact of nonsynonymous single-nucleotide polymorphisms on the protein stability of thiopurine S-methyltransferase as well as ATP-binding cassette (ABC) transporters including ABCC4, cystic fibrosis transmembrane conductance regulator (CFTR, ABCC7), ABCC11, and ABCG2. Furthermore, we will discuss the molecular mechanisms underlying posttranslational modifications (intramolecular and intermolecular disulfide bond formation and N-linked glycosylation) and ubiquitin-mediated proteasomal degradation of ABCG2, one of the major drug transporter proteins in humans.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
155 Effect of Mutations and Nonsynonymous SNPs on Protein Trafficking, Maturation, or ERAD of ABC Transporters Protein AA Mutation/SNP Effect on Protein Reference ABCA1 W590S Mutation Functional defect 115 R587W Mutation Impaired glycol processing 115 Q597R Mutation Impaired glycol processing, ERAD 115,116 Y1532C Mutation Altered protein trafficking 117 R1925Q Mutation Altered protein trafficking 118 ABCA3 R43L Mutation Altered protein trafficking 119 L101P Mutation Altered protein trafficking 119 R280C Mutation Altered protein trafficking 119 ABCA4 L541P Mutation Mislocalization 120 R602W Mutation Mislocalization 120 A1038V Mutation Mislocalization 120 C1490Y Mutation Mislocalization 120 ABCB1a G268V Mutation ERAD 121 G341C Mutation ERAD 121 I1196S Mutation Reduced glycosylation 122 ABCB4 I541F Mutation Accumulation in ER 123 ABCB11a E135K Mutation Reduced level of mature protein 124 L198P Mutation Reduced level of mature protein 124 E297G Mutation Reduced level of mature protein 124 L413W Mutation Reduced level of mature protein 124 R432T Mutation Reduced level of mature protein 124 D482G Mutation Immature protein in ER 124,125 N490D Mutation Reduced level of mature protein 124 A570T Mutation Reduced level of mature protein 124 T655I Mutation Reduced level of mature protein 124 Y818F SNP Moderate reduction of protein 124 G982R Mutation Retention in ER 125 R1153C Mutation ERAD 125 R1286Q Mutation Retention in ER 125 ABCC2a R768W Mutation Impaired protein trafficking 126 I1173F Mutation Impaired protein maturation 127 R1392 Mutation Impaired protein maturation 128 M1393 Mutation Impaired protein maturation 129 ABCC4a E757K SNP Altered protein trafficking 23 ABCC7 F508 Mutation Misfolding, ERAD 36-39,130 G85E Mutation Impaired protein maturation 130-132 G91R Mutation Impaired protein maturation 130-132 N1303K Mutation Impaired protein maturation 130-132 ABCC8 WT Wild type Ubiquitin-proteasome degradation 133 A116P Mutation Ubiquitin-proteasome degradation 133 V187D Mutation Ubiquitin-proteasome degradation 133 F1388 Mutation Impaired protein trafficking 134 L1544P Mutation Impaired protein trafficking 135,136 ABCC11a G180R SNP Ubiquitin-proteasome degradation 50 27 Mutation Ubiquitin-proteasome degradation 50 ABCG2a V12M SNP Altered protein localization 96 Q141K SNP Ubiquitin-proteasome degradation 102 F208S SNP Ubiquitin-proteasome degradation 78,99 S441N SNP Ubiquitin-proteasome degradation 78,99 Mutations of ABCA1, ABCA3, ABCA4, ABCB4, ABCB11, ABCC2, ABCC7 (CFTR), and ABCC8 are associated with Tangier disease, fatal surfactant deficiency, Stargardt disease, progressive familial intrahepatic cholestasis type 3 (PFIC-3), progressive familial intrahepatic cholestasis type 2 (PFIC-2), Dubin-Johnson syndrome, cystic fibrosis, and familial hyperinsulinism, respectively. Login to comment

[hide] CFTR: mechanism of anion conduction. Physiol Rev. 1999 Jan;79(1 Suppl):S47-75. Dawson DC, Smith SS, Mansoura MK
CFTR: mechanism of anion conduction.
Physiol Rev. 1999 Jan;79(1 Suppl):S47-75., [PMID:9922376]

Abstract [show]
CFTR: Mechanism of Anion Conduction. Physiol. Rev. 79, Suppl.: S47-S75, 1999. - The purpose of this review is to collect together the results of recent investigations of anion conductance by the cystic fibrosis transmembrane conductance regulator along with some of the basic background that is a prerequisite for developing some physical picture of the conduction process. The review begins with an introduction to the concepts of permeability and conductance and the Nernst-Planck and rate theory models that are used to interpret these parameters. Some of the physical forces that impinge on anion conductance are considered in the context of permeability selectivity and anion binding to proteins. Probes of the conduction process are considered, particularly permeant anions that bind tightly within the pore and block anion flow. Finally, structure-function studies are reviewed in the context of some predictions for the origin of pore properties.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
590 In addi-lectivity (see sect. IV), and in view of the roles of arginines in anion binding in other proteins (see sect. VE2), it is tion, Mansoura et al. (101) found that neutral (G91A), acidic (G91E), and basic (G91R) substitutions in this TMtempting to suggest that this TM might actually line the pore as suggested by the voltage-dependent accessibility had no effect on SCN binding or the sensitivity of the constructs to activation by IBMX, although the shape ofof some TM6 residues to MTS reagents (see sect. VF). Login to comment
591 Linsdell and Hanrahan (92, 93) suggested that the cyto- the i-V plot was altered for G91R. Login to comment

[hide] Membrane-integration characteristics of two ABC tr... J Mol Biol. 2009 Apr 17;387(5):1153-64. Epub 2009 Feb 21. Enquist K, Fransson M, Boekel C, Bengtsson I, Geiger K, Lang L, Pettersson A, Johansson S, von Heijne G, Nilsson I
Membrane-integration characteristics of two ABC transporters, CFTR and P-glycoprotein.
J Mol Biol. 2009 Apr 17;387(5):1153-64. Epub 2009 Feb 21., [PMID:19236881]

Abstract [show]
To what extent do corresponding transmembrane helices in related integral membrane proteins have different membrane-insertion characteristics? Here, we compare, side-by-side, the membrane insertion characteristics of the 12 transmembrane helices in the adenosine triphosphate-binding cassette (ABC) transporters, P-glycoprotein (P-gp) and the cystic fibrosis transmembrane conductance regulator (CFTR). Our results show that 10 of the 12 CFTR transmembrane segments can insert independently into the ER membrane. In contrast, only three of the P-gp transmembrane segments are independently stable in the membrane, while the majority depend on the presence of neighboring loops and/or transmembrane segments for efficient insertion. Membrane-insertion characteristics can thus vary widely between related proteins.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
113 For CFTR, we chose mutations located in TM1CFTR (F87L, G91R), TM3CFTR (P205S, L206W), TM4CFTR (C225R), TM5CFTR (DF311, G314E), TM6CFTR (R334L/W, I336K/R/D, I340N/S, L346P, R347L/H), TM8CFTR (S909I, S912L), TM9CFTR (I1005R, A1006E), TM10CFTR (Y1032N), and TM12CFTR (M1137R, ΔM1140, M1140K), or close to the TM region of TM1CFTR (R74W, L102R/P), TMF2CFTR (R117P/L, L137P), and TM11CFTR (M1101K/R). Login to comment
264 Structural cues involved in endoplasmic reticulum degradation of G85E and G91R mutant cystic fibrosis transmembrane conductance regulator. J. Clin. Login to comment
109 For CFTR, we chose mutations located in TM1CFTR (F87L, G91R), TM3CFTR (P205S, L206W), TM4CFTR (C225R), TM5CFTR (DF311, G314E), TM6CFTR (R334L/W, I336K/R/D, I340N/S, L346P, R347L/H), TM8CFTR (S909I, S912L), TM9CFTR (I1005R, A1006E), TM10CFTR (Y1032N), and TM12CFTR (M1137R, ƊM1140, M1140K), or close to the TM region of TM1CFTR (R74W, L102R/P), TMF2CFTR (R117P/L, L137P), and TM11CFTR (M1101K/R). Login to comment
261 Structural cues involved in endoplasmic reticulum degradation of G85E and G91R mutant cystic fibrosis transmembrane conductance regulator. J. Clin. Login to comment

[hide] Arsenite regulates Cystic Fibrosis Transmembrane C... Cell Physiol Biochem. 2005;16(1-3):109-18. Maitra R, Hamilton JW
Arsenite regulates Cystic Fibrosis Transmembrane Conductance Regulator and P-glycoprotein: evidence of pathway independence.
Cell Physiol Biochem. 2005;16(1-3):109-18., [PMID:16121039]

Abstract [show]
In the past, people have argued for and against the theory of reciprocal regulation of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) and P-glycoprotein (Pgp). Data have indicated that this may occur in vitro during drug-induced selection of cells, and in vivo during development. Much of this debate has been caused by a severe lack of mechanistic details involved in such regulation. Our past data indicate that certain Pgp modulators can affect CFTR expression and function. The goal of this study was to investigate the effects of trivalent arsenic (arsenite), a known transcriptional activator of Pgp, on CFTR expression. In vitro analyses in T-84 cells that express basal levels of Pgp and CFTR were conducted using a variety of molecular techniques. Expressions of both genes were altered following treatment with arsenite in a dose- and time-dependent fashion. CFTR expression was suppressed almost three-fold by arsenite, along with a concomitant increase in P-glycoprotein expression. We also report that a member of the MAPK-family, the ERK-mediated signaling cascade is implicated in suppression of CFTR expression following treatment with arsenite. However, this particular pathway is not involved in regulation of P-glycoprotein expression in T-84 cells following treatment with arsenite. Thus, the regulatory pathways that control functional expression of CFTR and P-glycoprotein following arsenite treatment in T-84 cells are distinct and independent.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
234 J Biol Chem 2000;275:24970-24976 19 Xiong X, Bragin A, Widdicombe JH, Cohn J, Skach WR: Structural cues involved in endoplasmic reticulum degradation of G85E and G91R mutant cystic fibrosis transmembrane conductance regulator. Login to comment

[hide] Alteration of CFTR transmembrane span integration ... Mol Biol Cell. 2011 Dec;22(23):4461-71. Epub 2011 Oct 12. Patrick AE, Karamyshev AL, Millen L, Thomas PJ
Alteration of CFTR transmembrane span integration by disease-causing mutations.
Mol Biol Cell. 2011 Dec;22(23):4461-71. Epub 2011 Oct 12., [PMID:21998193]

Abstract [show]
Many missense mutations in the cystic fibrosis transmembrane conductance regulator protein (CFTR) result in its misfolding, endoplasmic reticulum (ER) accumulation, and, thus, cystic fibrosis. A number of these mutations are located in the predicted CFTR transmembrane (TM) spans and have been projected to alter span integration. However, the boundaries of the spans have not been precisely defined experimentally. In this study, the ER luminal integration profiles of TM1 and TM2 were determined using the ER glycosylation machinery, and the effects of the CF-causing mutations G85E and G91R thereon were assessed. The mutations either destabilize the integrated conformation or alter the TM1 ER integration profile. G85E misfolding is based in TM1 destabilization by glutamic acid and loss of glycine and correlates with the temperature-insensitive ER accumulation of immature full-length CFTR harboring the mutation. By contrast, temperature-dependent misfolding owing to the G91R mutation depends on the introduction of the basic side chain rather than the loss of the glycine. This work demonstrates that CF-causing mutations predicted to have similar effects on CFTR structure actually result in disparate molecular perturbations that underlie ER accumulation and the pathology of CF.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
3 In this study, the ER luminal integration profiles of TM1 and TM2 were determined using the ER glycosylation machinery, and the effects of the CF-causing mutations G85E and G91R thereon were assessed. Login to comment
6 By contrast, temperature-dependent misfolding owing to the G91R mutation depends on the introduction of the basic side chain rather than the loss of the glycine. Login to comment
36 The CF-causing mutants G91R and G85E are in the original predicted TM1 span, residues 81-102 (Riordan et al., 1989). Login to comment
41 Consistent with reduced stability, full-length G91R CFTR accumulates in the ER, and multiple domains exhibit increased proteolytic susceptibility in mammalian cells (Du and Lukacs, 2009). Login to comment
66 In this study, the span boundaries or integration profiles of TM1 and TM2 were determined and the effects of the CF-causing mutations G85E and G91R assessed. Login to comment
67 The G91R and G85E mutations Figure 1: Predicted TM1 and TM2 spans. Login to comment
69 The positions of the CF-causing mutants G85E and G91R are indicated by triangles. Login to comment
71 This work further elucidates CFTR membrane-spanning structures and provides mechanistic insight into the molecular pathology of the G85E and G91R CF-causing mutations. Login to comment
76 Moreover, when the CF-causing mutations G85E and G91R were analyzed using these algorithms, variable effects of mutations were predicted, including shortening, no affect, no TM, or shifting TM1 boundaries (Supplemental Table S1). Login to comment
114 Core glycosylation analysis of WT, G91R, and G85E CFTR containing the artificial ECL1 site and lacking the ECL4 sites was performed by deletion of residues between the glycosylation site and TM1 (A) or between the glycosylation site and TM2 (B). Login to comment
117 Schematics of the experimentally identified ER integration profiles in WT (C) or G91R (D) and G85E (E) mutants. Login to comment
130 Determining the ER integration profiles of CF-causing mutants To assess effects of the CF-causing mutants G91R and G85E on the TM1 ER integration profile, these mutations were analyzed using the ECL1 site core glycosylation assay. Login to comment
131 The G85E and G91R mutations introduce an ionizable group into or near the predicted TM1 span and might be reasonably expected to alter its ER integration profile (Xiong et al., 1997). Login to comment
133 The G91R and G85E mutations in CFTR containing the natural ECL4 sites or the ECL1 site resulted in misfolding and accumulation in the ER (Supplemental Figure S3). Login to comment
136 Thus, in the same manner as WT ECL1, glycosylation analysis was used to characterize the TM1 integration profiles for the G91R and G85E mutants. Login to comment
137 Cystic fibrosis-causing mutant G91R shifts the ER integrated profile of TM1 The effect of the G91R mutation on the TM1 ER integrated profile was tested by its introduction into the ECL1 core glycosylation assay. Login to comment
139 By contrast, G91R N-terminal deletion constructs were completely core glycosylated until a 14-residue deletion resulted in partial core glycosylation, and a 16-residue deletion was completely nonglycosylated (Figure 4A), indicating the edge of the mutant TM integration profile shifted by two or three residues. Login to comment
140 Thus, instead of I86 as in the wild type, L88 is 12 residues from the ECL1 site in the mutant, positioning it at the G91R TM1 ER luminal integration profile edge (Figure 4D). Login to comment
148 As is the case for G91R, the L88 TM1 integration profile edge is slightly shifted from WT. Login to comment
163 The reaction of C76 with AMS in the presence of G91R and G85E was also tested. Login to comment
164 In WT and G91R, no shift is observed, indicating that position 76 is protected by the membrane (Figure 5, B and C). Login to comment
166 This result is consistent with glycosylation scanning results and an initial positioning of TM1 in the presence of G85E that is more C-terminal than for either WT or G91R. Login to comment
168 Role of the ionizable side chain in altered G91R and G85E TM1 ER integration profiles In the G91R and G85E mutants, an ionizable side chain replaces the glycine Cα hydrogen. Login to comment
174 This integration profile is the same as the G91R TM1integration profile and one of the two extreme G85E TM1 integration profiles. Login to comment
176 Role of the ionizable side chain in trafficking of G91R and G85E The data from the glycosylation assay demonstrate that the G85E mutant splits the integration profile of TM1, whereas the G91R, G85A, and G91A mutants do not. Login to comment
179 G91A is both core and complex glycosylated and traffics to the cell surface, indicating that introduction of arginine rather than loss of glycine causes G91R ER accumulation. Login to comment
186 At the lower temperature, both G91R and G85A mutants partially traffic from the ER and are thus temperature sensitive, similar to ΔF508. Login to comment
194 This study determined the TM1 and TM2 ER luminal integration profile edges and CF-causing mutant G91R and G85E effects on TM1, using the mammalian ER luminal core glycosylation machinery. Login to comment
204 (B) A CFTR construct containing C76 with G91R or G85E was exposed to AMS and examined for the presence of an interaction by gel shift. Login to comment
206 (C) Schematics of G91R and G85E mutant CFTR three-TM constructs with the relative positions of C76 and the mutants labeled. Login to comment
235 HeLa cell trafficking of WT, ΔF508, G85E, G85A, G91R, and G91A mutant CFTR at 37°C was analyzed by Western blot analysis (A). Login to comment
249 Thus CF mutations, such as G91R, in the C-terminal region may be within or exposed to the ER lumen during translation and integration. Login to comment
250 Consistent with this is the modest shift in the G91R-CFTR TM1 ER integration profile. Login to comment
251 The data here and in previous reports indicate that G91R, but not G91A, disrupts CFTR trafficking in the cell and has significant effects on the stability and assembly of full-length CFTR (Xiong et al., 1997; Younger et al., 2006; Rosser et al., 2008; Du and Lukacs, 2009). Login to comment
270 Previous work demonstrated that the G85E and G91R mutations also disrupt later steps in CFTR folding, particularly interdomain interactions, which were proposed to underlie mutant recognition by ER quality control machinery (Xiong et al., 1997). Login to comment
273 The G91R mutant was predicted to have a similar effect on CFTR (Xiong et al., 1997). Login to comment
274 Yet the experimental evidence presented here distinguishes G91R from G85E with respect to perturbations from the ionizable side chain, the role of glycine, and temperature sensitivity. Login to comment
277 It is striking that the corrector compound 4 exhibited mutant-specific effects, partially rescuing the G91R but not G85E CFTR (Grove et al., 2009). Login to comment
286 The mutations G91R, G91A, G85E, and G85A were introduced into CFTR constructs containing the natural, artificial, and deletion mutants on the artificial site. Login to comment

[hide] Sequential quality-control checkpoints triage misf... Cell. 2006 Aug 11;126(3):571-82. Younger JM, Chen L, Ren HY, Rosser MF, Turnbull EL, Fan CY, Patterson C, Cyr DM
Sequential quality-control checkpoints triage misfolded cystic fibrosis transmembrane conductance regulator.
Cell. 2006 Aug 11;126(3):571-82., [PMID:16901789]

Abstract [show]
Cystic fibrosis arises from the misfolding and premature degradation of CFTR Delta F508, a Cl- ion channel with a single amino acid deletion. Yet, the quality-control machinery that selects CFTR Delta F508 for degradation and the mechanism for its misfolding are not well defined. We identified an ER membrane-associated ubiquitin ligase complex containing the E3 RMA1, the E2 Ubc6e, and Derlin-1 that cooperates with the cytosolic Hsc70/CHIP E3 complex to triage CFTR and CFTR Delta F508. Derlin-1 serves to retain CFTR in the ER membrane and interacts with RMA1 and Ubc6e to promote CFTR's proteasomal degradation. RMA1 is capable of recognizing folding defects in CFTR Delta F508 coincident with translation, whereas the CHIP E3 appears to act posttranslationally. A folding defect in CFTR Delta F508 detected by RMA1 involves the inability of CFTR's second membrane-spanning domain to productively interact with amino-terminal domains. Thus, the RMA1 and CHIP E3 ubiquitin ligases act sequentially in ER membrane and cytosol to monitor the folding status of CFTR and CFTR Delta F508.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
207 Hence, we compared the ability of RMA1 and CHIP to sense the folding defect caused by the G91R mutation in transmembrane helix I, which prevents proper assembly of CFTR (Xiong et al., 1997). Login to comment
208 Indeed, CFTR1162X G91R was found to be sensitive to elevation of RMA1 activity, but not to CHIP. Login to comment
221 First, the elevation of RMA1 activity drives the degradation of CFTR biogenic intermediates that have the G91R mutation in MSDI, which is degraded by ERAD due to defects in MSD assembly (Xiong et al., 1997). Login to comment

[hide] Genotyping microarray for the detection of more th... J Mol Diagn. 2005 Aug;7(3):375-87. Schrijver I, Oitmaa E, Metspalu A, Gardner P
Genotyping microarray for the detection of more than 200 CFTR mutations in ethnically diverse populations.
J Mol Diagn. 2005 Aug;7(3):375-87., [PMID:16049310]

Abstract [show]
Cystic fibrosis (CF), which is due to mutations in the cystic fibrosis transmembrane conductance regulator gene, is a common life-shortening disease. Although CF occurs with the highest incidence in Caucasians, it also occurs in other ethnicities with variable frequency. Recent national guidelines suggest that all couples contemplating pregnancy should be informed of molecular screening for CF carrier status for purposes of genetic counseling. Commercially available CF carrier screening panels offer a limited panel of mutations, however, making them insufficiently sensitive for certain groups within an ethnically diverse population. This discrepancy is even more pronounced when such carrier screening panels are used for diagnostic purposes. By means of arrayed primer extension technology, we have designed a genotyping microarray with 204 probe sites for CF transmembrane conductance regulator gene mutation detection. The arrayed primer extension array, based on a platform technology for disease detection with multiple applications, is a robust, cost-effective, and easily modifiable assay suitable for CF carrier screening and disease detection.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
51 Complete List of Mutations Detectable with the CF APEX Assay CFTR location Amino acid change Nucleotide change 1 E 1 Frameshift 175delC 2 E 2,3 Frameshift del E2, E3 3 E 2 W19C 189 GϾT 4 E 2 Q39X 247 CϾT 5 IVS 2 Possible splicing defect 296 ϩ 12 TϾC 6 E 3 Frameshift 359insT 7 E 3 Frameshift 394delTT 8 E 3 W57X (TAG) 302GϾA 9 E 3 W57X (TGA) 303GϾA 10 E 3 E60X 310GϾT 11 E 3 P67L 332CϾT 12 E 3 R74Q 353GϾA 13 E 3 R75X 355CϾT 14 E 3 G85E 386GϾA 15 E 3 G91R 403GϾA 16 IVS 3 Splicing defect 405 ϩ 1GϾA 17 IVS 3 Possible splicing defect 405 ϩ 3AϾC 18 IVS 3 Splicing defect 406 - 1GϾA 19 E 4 E92X 406GϾT 20 E 4 E92K 406GϾA 21 E 4 Q98R 425AϾG 22 E 4 Q98P 425AϾC 23 E 4 Frameshift 444delA 24 E 4 Frameshift 457TATϾG 25 E 4 R117C 481CϾT 26 E 4 R117H 482GϾA 27 E 4 R117P 482GϾC 28 E 4 R117L 482GϾT 29 E 4 Y122X 498TϾA 30 E 4 Frameshift 574delA 31 E 4 I148T 575TϾC 32 E 4 Splicing defect 621GϾA 33 IVS 4 Splicing defect 621 ϩ 1GϾT 34 IVS 4 Splicing defect 621 ϩ 3AϾG 35 E 5 Frameshift 624delT 36 E 5 Frameshift 663delT 37 E 5 G178R 664GϾA 38 E 5 Q179K 667CϾA 39 IVS 5 Splicing defect 711 ϩ 1GϾT 40 IVS 5 Splicing defect 711 ϩ 1GϾA 41 IVS 5 Splicing defect 712 - 1GϾT 42 E 6a H199Y 727CϾT 43 E 6a P205S 745CϾT 44 E 6a L206W 749TϾG 45 E 6a Q220X 790CϾT 46 E 6b Frameshift 935delA 47 E 6b Frameshift 936delTA 48 E 6b N287Y 991AϾT 49 IVS 6b Splicing defect 1002 - 3TϾG 50 E 7 ⌬F311 3-bp del between nucleotides 1059 and 1069 51 E 7 Frameshift 1078delT 52 E 7 Frameshift 1119delA 53 E 7 G330X 1120GϾT 54 E 7 R334W 1132CϾT 55 E 7 I336K 1139TϾA 56 E 7 T338I 1145CϾT 57 E 7 Frameshift 1154insTC 58 E 7 Frameshift 1161delC 59 E 7 L346P 1169TϾC 60 E 7 R347H 1172GϾA 61 E 7 R347P 1172GϾC 62 E 7 R347L 1172GϾT 63 E 7 R352Q 1187GϾA 64 E 7 Q359K/T360K 1207CϾA and 1211CϾA 65 E 7 S364P 1222TϾC 66 E 8 Frameshift 1259insA 67 E 8 W401X (TAG) 1334GϾA 68 E 8 W401X (TGA) 1335GϾA 69 IVS 8 Splicing changes 1342 - 6 poly(T) variants 5T/7T/9T 70 IVS 8 Splicing defect 1342 - 2AϾC Table 1. Continued CFTR location Amino acid change Nucleotide change 71 E 9 A455E 1496CϾA 72 E 9 Frameshift 1504delG 73 E 10 G480C 1570GϾT 74 E 10 Q493X 1609CϾT 75 E 10 Frameshift 1609delCA 76 E 10 ⌬I507 3-bp del between nucleotides 1648 and 1653 77 E 10 ⌬F508 3-bp del between nucleotides 1652 and 1655 78 E 10 Frameshift 1677delTA 79 E 10 V520F 1690GϾT 80 E 10 C524X 1704CϾA 81 IVS 10 Possible splicing defect 1717 - 8GϾA 82 IVS 10 Splicing defect 1717 - 1GϾA 83 E 11 G542X 1756GϾT 84 E 11 G551D 1784GϾA 85 E 11 Frameshift 1784delG 86 E 11 S549R (AϾC) 1777AϾC 87 E 11 S549I 1778GϾT 88 E 11 S549N 1778GϾA 89 E 11 S549R (TϾG) 1779TϾG 90 E 11 Q552X 1786CϾT 91 E 11 R553X 1789CϾT 92 E 11 R553G 1789CϾG 93 E 11 R553Q 1790GϾA 94 E 11 L558S 1805TϾC 95 E 11 A559T 1807GϾA 96 E 11 R560T 1811GϾC 97 E 11 R560K 1811GϾA 98 IVS 11 Splicing defect 1811 ϩ 1.6 kb AϾG 99 IVS 11 Splicing defect 1812 - 1GϾA 100 E 12 Y563D 1819TϾG 101 E 12 Y563N 1819TϾA 102 E 12 Frameshift 1833delT 103 E 12 D572N 1846GϾA 104 E 12 P574H 1853CϾA 105 E 12 T582R 1877CϾG 106 E 12 E585X 1885GϾT 107 IVS 12 Splicing defect 1898 ϩ 5GϾT 108 IVS 12 Splicing defect 1898 ϩ 1GϾA 109 IVS 12 Splicing defect 1898 ϩ 1GϾC 110 IVS 12 Splicing defect 1898 ϩ 1GϾT 111 E 13 Frameshift 1924del7 112 E 13 del of 28 amino acids 1949del84 113 E 13 I618T 1985TϾC 114 E 13 Frameshift 2183AAϾG 115 E 13 Frameshift 2043delG 116 E 13 Frameshift 2055del9ϾA 117 E 13 D648V 2075TϾA 118 E 13 Frameshift 2105-2117 del13insAGAA 119 E 13 Frameshift 2108delA 120 E 13 R668C 2134CϾT 121 E 13 Frameshift 2143delT 122 E 13 Frameshift 2176insC 123 E 13 Frameshift 2184delA 124 E 13 Frameshift 2184insA 125 E 13 Q685X 2185CϾT 126 E 13 R709X 2257CϾT 127 E 13 K710X 2260AϾT 128 E 13 Frameshift 2307insA 129 E 13 V754M 2392GϾA 130 E 13 R764X 2422CϾT 131 E 14a W846X 2670GϾA 132 E 14a Frameshift 2734delGinsAT 133 E 14b Frameshift 2766del8 134 IVS 14b Splicing defect 2789 ϩ 5GϾA 135 IVS 14b Splicing defect 2790 - 2AϾG 136 E 15 Q890X 2800CϾT 137 E 15 Frameshift 2869insG 138 E 15 S945L 2966CϾT 139 E 15 Frameshift 2991del32 140 E 16 Splicing defect 3120GϾA interrogation: ACCAACATGTTTTCTTTGATCTTAC 3121-2A3G,T S; 5Ј-ACCAACATGTTTTCTTTGATCTTAC A GTTGTTATTAATTGTGATTGGAGCTATAG-3Ј; CAACAA- TAATTAACACTAACCTCGA 3121-2A3G,T AS. Login to comment
150 Primers Generated to Create Synthetic Templates That Serve As Positive Mutation Controls Primer name Sense strand 5Ј 3 3Ј Name Antisense strand 5Ј 3 3Ј 175delC synt F T(15)ATTTTTTTCAGGTGAGAAGGTGGCCA 175delC synt R T(15)ATTTGGAGACAACGCTGGCCTTTTCC W19C synt F T(15)TACCAGACCAATTTTGAGGAAAGGAT W19C synt R T(15)ACAGCTAAAATAAAGAGAGGAGGAAC Q39X synt F T(15)TAAATCCCTTCTGTTGATTCTGCTGA Q39X synt R T(15)AGTATATGTCTGACAATTCCAGGCGC 296 ϩ 12TϾC synt F T(15)CACATTGTTTAGTTGAAGAGAGAAAT 296 ϩ 12TϾC synt R T(15)GCATGAACATACCTTTCCAATTTTTC 359insT synt F T(15)TTTTTTTCTGGAGATTTATGTTCTAT 359insT synt R T(15)AAAAAAACATCGCCGAAGGGCATTAA E60X synt F T(15)TAGCTGGCTTCAAAGAAAAATCCTAA E60X synt R T(15)ATCTATCCCATTCTCTGCAAAAGAAT P67L synt F T(15)TTAAACTCATTAATGCCCTTCGGCGA P67L synt R T(15)AGATTTTTCTTTGAAGCCAGCTCTCT R74Q synt F T(15)AGCGATGTTTTTTCTGGAGATTTATG R74Q synt R T(15)TGAAGGGCATTAATGAGTTTAGGATT R75X synt F T(15)TGATGTTTTTTCTGGAGATTTATGTT R75X synt R T(15)ACCGAAGGGCATTAATGAGTTTAGGA W57X(TAG) synt F T(15)AGGATAGAGAGCTGGCTTCAAAGAAA W57X(TAG) synt R T(15)TATTCTCTGCAAAAGAATAAAAAGTG W57X(TGA) synt F T(15)AGATAGAGAGCTGGCTTCAAAGAAAA W57X(TGA) synt R T(15)TCATTCTCTGCAAAAGAATAAAAAGT G91R synt F T(15)AGGGTAAGGATCTCATTTGTACATTC G91R synt R T(15)TTAAATATAAAAAGATTCCATAGAAC 405 ϩ 1GϾA synt F T(15)ATAAGGATCTCATTTGTACATTCATT 405 ϩ 1GϾA synt R T(15)TCCCTAAATATAAAAAGATTCCATAG 405 ϩ 3AϾC synt F T(15)CAGGATCTCATTTGTACATTCATTAT 405 ϩ 3AϾC synt R T(15)GACCCCTAAATATAAAAAGATTCCAT 406 - 1GϾA synt F T(15)AGAAGTCACCAAAGCAGTACAGCCTC 406 - 1GϾA synt R T(15)TTACAAAAGGGGAAAAACAGAGAAAT E92X synt F T(15)TAAGTCACCAAAGCAGTACAGCCTCT E92X synt R T(15)ACTACAAAAGGGGAAAAACAGAGAAA E92K synt F T(15)AAAGTCACCAAAGCAGTACAGCCTCT E92K synt R T(15)TCTACAAAAGGGGAAAAACAGAGAAA 444delA synt F T(15)GATCATAGCTTCCTATGACCCGGATA 444delA synt R T(15)ATCTTCCCAGTAAGAGAGGCTGTACT 574delA synt F T(15)CTTGGAATGCAGATGAGAATAGCTAT 574delA synt R T(15)AGTGATGAAGGCCAAAAATGGCTGGG 621GϾA synt F T(15)AGTAATACTTCCTTGCACAGGCCCCA 621GϾA synt R T(15)TTTCTTATAAATCAAACTAAACATAG Q98P synt F T(15)CGCCTCTCTTACTGGGAAGAATCATA Q98P synt R T(15)GGTACTGCTTTGGTGACTTCCTACAA 457TATϾG synt F T(15)GGACCCGGATAACAAGGAGGAACGCT 457TATϾG synt R T(15)CGGAAGCTATGATTCTTCCCAGTAAG I148T synt F T(15)CTGGAATGCAGATGAGAATAGCTATG I148T synt R T(15)GTGTGATGAAGGCCAAAAATGGCTGG 624delT synt F T(15)CTTAAAGCTGTCAAGCCGTGTTCTAG 624delT synt R T(15)TAAGTCTAAAAGAAAAATGGAAAGTT 663delT synt F T(15)ATGGACAACTTGTTAGTCTCCTTTCC 663delT synt R T(15)CATACTTATTTTATCTAGAACACGGC G178R synt F T(15)AGACAACTTGTTAGTCTCCTTTCCAA G178R synt R T(15)TAATACTTATTTTATCTAGAACACGG Q179K synt F T(15)AAACTTGTTAGTCTCCTTTCCAACAA Q179K synt R T(15)TTCCAATACTTATTTTATCTAGAACA 711 ϩ 5GϾA synt F T(15)ATACCTATTGATTTAATCTTTTAGGC 711 ϩ 5GϾA synt R T(15)TTATACTTCATCAAATTTGTTCAGGT 712 - 1GϾT synt F T(15)TGGACTTGCATTGGCACATTTCGTGT 712 - 1GϾT synt R T(15)TATGGAAAATAAAAGCACAGCAAAAAC H199Y synt F T(15)TATTTCGTGTGGATCGCTCCTTTGCA H199Y synt R T(15)TATGCCAATGCTAGTCCCTGGAAAATA P205S synt F T(15)TCTTTGCAAGTGGCACTCCTCATGGG P205S synt R T(15)TAAGCGATCCACACGAAATGTGCCAAT L206W synt F T(15)GGCAAGTGGCACTCCTCATGGGGCTA L206W synt R T(15)TCAAGGAGCGATCCACACGAAATGTGC Q220X synt F T(15)TAGGCGTCTGCTTTCTGTGGACTTGG Q220X synt R T(15)TATAACAACTCCCAGATTAGCCCCATG 936delTA synt F T(15)AATCCAATCTGTTAAGGCATACTGCT 936delTA synt R T(15)TGATTTTCAATCATTTCTGAGGTAATC 935delA synt F T(15)GAAATATCCAATCTGTTAAGGCATAC 935delA synt R T(15)TATTTCAATCATTTCTGAGGTAATCAC N287Y synt F T(15)TACTTAAGACAGTAAGTTGTTCCAAT N287Y synt R T(15)TATTCAATCATTTTTTCCATTGCTTCT 1002 - 3TϾG synt F T(15)GAGAACAGAACTGAAACTGACTCGGA 1002 - 3TϾG synt R T(15)TCTAAAAAACAATAACAATAAAATTCA 1154insTC syntwt F T(15)ATCTCATTCTGCATTGTTCTGCGCAT 1154insTC syntwt R T(15)TTGAGATGGTGGTGAATATTTTCCGGA 1154insTC syntmt F T(15)TCTCTCATTCTGCATTGTTCTGCGCAT 1154insTC syntmt R T(15)TAGAGATGGTGGTGAATATTTTCCGGA DF311 mt syntV1 F T(15)CCTTCTTCTCAGGGTTCTTTGTGGTG dF311 mt syntV1 R T(15)GAGAAGAAGGCTGAGCTATTGAAGTATC G330X synt F T(15)TGAATCATCCTCCGGAAAATATTCAC G330X synt R T(15)ATTTGATTAGTGCATAGGGAAGCACA S364P synt F T(15)CCTCTTGGAGCAATAAACAAAATACA S364P synt R T(15)GGTCATACCATGTTTGTACAGCCCAG Q359K/T360K mt synt F T(15)AAAAAATGGTATGACTCTCTTGGAGC Q359K/T360K mt synt R T(15)TTTTTTACAGCCCAGGGAAATTGCCG 1078delT synt F T(15)CTTGTGGTGTTTTTATCTGTGCTTCC 1078delT synt R T(15)CAAGAACCCTGAGAAGAAGAAGGCTG 1119delA synt F T(15)CAAGGAATCATCCTCCGGAAAATATT 1119delA synt R T(15)CTTGATTAGTGCATAGGGAAGCACAG 1161delC synt F T(15)GATTGTTCTGCGCATGGCGGTCACTC 1161delC synt R T(15)TCAGAATGAGATGGTGGTGAATATTT T338I synt F T(15)TCACCATCTCATTCTGCATTGTTCTG T338I synt R T(15)ATGAATATTTTCCGGAGGATGATTCC R352Q synt F T(15)AGCAATTTCCCTGGGCTGTACAAACA R352Q synt R T(15)TGAGTGACCGCCATGCGCAGAACAAT L346P synt F T(15)CGCGCATGGCGGTCACTCGGCAATTT L346P synt R T(15)GGAACAATGCAGAATGAGATGGTGGT 1259insA synt F T(15)AAAAAGCAAGAATATAAGACATTGGA 1259insA synt R T(15)TTTTTGTAAGAAATCCTATTTATAAA W401X(TAG)mtsynt F T(15)AGGAGGAGGTCAGAATTTTTAAAAAA W401X(TAG)mtsynt R T(15)TAGAAGGCTGTTACATTCTCCATCAC W401X(TGA) synt F T(15)AGAGGAGGTCAGAATTTTTAAAAAAT W401X(TGA) synt R T(15)TCAGAAGGCTGTTACATTCTCCATCA 1342 - 2AϾC synt F T(15)CGGGATTTGGGGAATTATTTGAGAAA 1342 - 2AϾC synt R T(15)GGTTAAAAAAACACACACACACACAC 1504delG synt F T(15)TGATCCACTGTAGCAGGCAAGGTAGT 1504delG synt R T(15)TCAGCAACCGCCAACAACTGTCCTCT G480C synt F T(15)TGTAAAATTAAGCACAGTGGAAGAAT G480C synt R T(15)ACTCTGAAGGCTCCAGTTCTCCCATA C524X synt F T(15)ACAACTAGAAGAGGTAAGAAACTATG C524X synt R T(15)TCATGCTTTGATGACGCTTCTGTATC V520F synt F T(15)TTCATCAAAGCAAGCCAACTAGAAGA V520F synt R T(15)AGCTTCTGTATCTATATTCATCATAG 1609delCA synt F T(15)TGTTTTCCTGGATTATGCCTGGCACC 1609delCA synt R T(15)CAGAACAGAATGAAATTCTTCCACTG 1717 - 8GϾA synt F T(15)AGTAATAGGACATCTCCAAGTTTGCA 1717 - 8GϾA synt R T(15)TAAAAATAGAAAATTAGAGAGTCACT 1784delG synt F T(15)AGTCAACGAGCAAGAATTTCTTTAGC 1784delG synt R T(15)ACTCCACTCAGTGTGATTCCACCTTC A559T synt F T(15)ACAAGGTGAATAACTAATTATTGGTC A559T synt R T(15)TTAAAGAAATTCTTGCTCGTTGACCT Q552X synt F T(15)TAACGAGCAAGAATTTCTTTAGCAAG Q552X synt R T(15)AACCTCCACTCAGTGTGATTCCACCT S549R(AϾC) synt F T(15)CGTGGAGGTCAACGAGCAAGAATTTC S549R(AϾC) synt R T(15)GCAGTGTGATTCTACCTTCTCCAAGA S549R(TϾG) synt F T(15)GGGAGGTCAACGAGCAAGTATTTC S549R(TϾG) synt R T(15)CCTCAGTGTGATTCCACCTTCTCCAA L558S synt F T(15)CAGCAAGGTGAATAACTAATTATTGG L558S synt R T(15)GAAGAAATTCTCGCTCGTTGACCTCC 1811 ϩ 1.6 kb AϾG synt F T(15)GTAAGTAAGGTTACTATCAATCACAC 1811 ϩ 1.6 kb AϾG synt R T(15)CATCTCAAGTACATAGGATTCTCTGT 1812 - 1GϾA synt F T(15)AAGCAGTATACAAAGATGCTGATTTG 1812 - 1GϾA synt R T(15)TTAAAAAGAAAATGGAAATTAAATTA D572N synt F T(15)AACTCTCCTTTTGGATACCTAGATGT D572N synt R T(15)TTAATAAATACAAATCAGCATCTTTG P574H synt F T(15)ATTTTGGATACCTAGATGTTTTAACA P574H synt R T(15)TGAGAGTCTAATAAATACAAATCAGC 1833delT synt F T(15)ATTGTATTTATTAGACTCTCCTTTTG 1833delT synt R T(15)CAATCAGCATCTTTGTATACTGCTCT Table 4. Continued Primer name Sense strand 5Ј 3 3Ј Name Antisense strand 5Ј 3 3Ј Y563D synt F T(15)GACAAAGATGCTGATTTGTATTTATT Y563D synt R T(15)CTACTGCTCTAAAAAGAAAATGGAAA T582R synt F T(15)GAGAAAAAGAAATATTTGAAAGGTAT T582R synt R T(15)CTTAAAACATCTAGGTATCCAAAAGG E585X synt F T(15)TAAATATTTGAAAGGTATGTTCTTTG E585X synt R T(15)ATTTTTCTGTTAAAACATCTAGGTAT 1898 ϩ 5GϾT synt F T(15)TTTCTTTGAATACCTTACTTATATTG 1898 ϩ 5GϾT synt R T(15)AATACCTTTCAAATATTTCTTTTTCT 1924del7 synt F T(15)CAGGATTTTGGTCACTTCTAAAATGG 1924del7 synt R T(15)CTGTTAGCCATCAGTTTACAGACACA 2055del9ϾA synt F T(15)ACATGGGATGTGATTCTTTCGACCAA 2055del9ϾA synt R T(15)TCTAAAGTCTGGCTGTAGATTTTGGA D648V synt F T(15)TTTCTTTCGACCAATTTAGTGCAGAA D648V synt R T(15)ACACATCCCATGAGTTTTGAGCTAAA K710X synt F T(15)TAATTTTCCATTGTGCAAAAGACTCC K710X synt R T(15)ATCGTATAGAGTTGATTGGATTGAGA I618T synt F T(15)CTTTGCATGAAGGTAGCAGCTATTTT I618T synt R T(15)GTTAATATTTTGTCAGCTTTCTTTAA R764X synt F T(15)TGAAGGAGGCAGTCTGTCCTGAACCT R764X synt R T(15)ATGCCTGAAGCGTGGGGCCAGTGCTG Q685X synt F T(15)TAATCTTTTAAACAGACTGGAGAGTT Q685X synt R T(15)ATTTTTTTGTTTCTGTCCAGGAGACA R709X synt F T(15)TGAAAATTTTCCATTGTGCAAAAGAC R709X synt R T(15)ATATAGAGTTGATTGGATTGAGAATA V754M synt F T(15)ATGATCAGCACTGGCCCCACGCTTCA V754M synt R T(15)TGCTGATGCGAGGCAGTATCGCCTCT 1949del84 synt F T(15)AAAAATCTACAGCCAGACTTTATCTC 1949del84 synt R T(15)TTTTTAGAAGTGACCAAAATCCTAGT 2108delA synt F T(15)GAATTCAATCCTAACTGAGACCTTAC 2108delA synt R T(15)ATTCTTCTTTCTGCACTAAATTGGTC 2176insC synt F T(15)CCAAAAAAACAATCTTTTAAACAGACTGGAGAG 2176insC synt R T(15)GGTTTCTGTCCAGGAGACAGGAGCAT 2184delA synt F T(15)CAAAAAACAATCTTTTAAACAGACTGG 2184delA synt R T(15)GTTTTTTGTTTCTGTCCAGGAGACAG 2105-2117 del13 synt F T(15)AAACTGAGACCTTACACCGTTTCTCA 2105-2117 del13 synt R T(15)TTTCTTTCTGCACTAAATTGGTCGAA 2307insA synt F T(15)AAAGAGGATTCTGATGAGCCTTTAGA 2307insA synt R T(15)TTTCGATGCCATTCATTTGTAAGGGA W846X synt F T(15)AAACACATACCTTCGATATATTACTGTCCAC W846X synt R T(15)TCATGTAGTCACTGCTGGTATGCTCT 2734G/AT synt F T(15)TTAATTTTTCTGGCAGAGGTAAGAAT 2734G/AT synt R T(15)TTAAGCACCAAATTAGCACAAAAATT 2766del8 synt F T(15)GGTGGCTCCTTGGAAAGTGAGTATTC 2766del8 synt R T(15)CACCAAAGAAGCAGCCACCTGGAATGG 2790 - 2AϾG synt F T(15)GGCACTCCTCTTCAAGACAAAGGGAA 2790 - 2AϾG synt R T(15)CGTAAAGCAAATAGGAAATCGTTAAT 2991del32 synt F T(15)TTCAACACGTCGAAAGCAGGTACTTT 2991del32 synt R T(15)AAACATTTTGTGGTGTAAAATTTTCG Q890X synt F T(15)TAAGACAAAGGGAATAGTACTCATAG Q890X synt R T(15)AAAGAGGAGTGCTGTAAAGCAAATAG 2869insG synt F T(15)GATTATGTGTTTTACATTTACGTGGG 2869insG synt R T(15)CACGAACTGGTGCTGGTGATAATCAC 3120GϾA synt F T(15)AGTATGTAAAAATAAGTACCGTTAAG 3120GϾA synt R T(15)TTGGATGAAGTCAAATATGGTAAGAG 3121 - 2AϾT synt F T(15)TGTTGTTATTAATTGTGATTGGAGCT 3121 - 2AϾT synt R T(15)AGTAAGATCAAAGAAAACATGTTGGT 3132delTG synt F T(15)TTGATTGGAGCCATAGCAGTTGTCGC 3132delTG synt R T(15)AATTAATAACAACTGTAAGATCAAAG 3271delGG synt F T(15)ATATGACAGTGAATGTGCGATACTCA 3271delGG synt R T(15)ATTCAGATTCCAGTTGTTTGAGTTGC 3171delC synt F T(15)ACCTACATCTTTGTTGCAACAGTGCC 3171delC synt R T(15)AGGTTGTAAAACTGCGACAACTGCTA 3171insC synt F T(15)CCCCTACATCTTTGTTGCTACAGTGC 3171insC synt R T(15)GGGGTTGTAAAACTGCGACAACTGCT 3199del6 synt F T(15)GAGTGGCTTTTATTATGTTGAGAGCATAT 3199del6 synt R T(15)CCACTGGCACTGTTGCAACAAAGATG M1101K synt F T(15)AGAGAATAGAAATGATTTTTGTCATC M1101K synt R T(15)TTTTGGAACCAGCGCAGTGTTGACAG G1061R synt F T(15)CGACTATGGACACTTCGTGCCTTCGG G1061R synt R T(15)GTTTTAAGCTTGTAACAAGATGAGTG R1066L synt F T(15)TTGCCTTCGGACGGCAGCCTTACTTT R1066L synt R T(15)AGAAGTGTCCATAGTCCTTTTAAGCT R1070P synt F T(15)CGCAGCCTTACTTTGAAACTCTGTTC R1070P synt R T(15)GGTCCGAAGGCACGAAGTGTCCATAG L1077P synt F T(15)CGTTCCACAAAGCTCTGAATTTACAT L1077P synt R T(15)GGAGTTTCAAAGTAAGGCTGCCGTCC W1089X synt F T(15)AGTTCTTGTACCTGTCAACACTGCGC W1089X synt R T(15)TAGTTGGCAGTATGTAAATTCAGAGC L1093P synt F T(15)CGTCAACACTGCGCTGGTTCCAAATG L1093P synt R T(15)GGGTACAAGAACCAGTTGGCAGTATG W1098R synt F T(15)CGGTTCCAAATGAGAATAGAAATGAT W1098R synt R T(15)GGCGCAGTGTTGACAGGTACAAGAAC Q1100P synt F T(15)CAATGAGAATAGAAATGATTTTTGTC Q1100P synt R T(15)GGGAACCAGCGCAGTGTTGACAGGTA D1152H synt F T(15)CATGTGGATAGCTTGGTAAGTCTTAT D1152H synt R T(15)GTATGCTGGAGTTTACAGCCCACTGC R1158X synt F T(15)TGATCTGTGAGCCGAGTCTTTAAGTT R1158X synt R T(15)ACATCTGAAATAAAAATAACAACATT S1196X synt F T(15)GACACGTGAAGAAAGATGACATCTGG S1196X synt R T(15)CAATTCTCAATAATCATAACTTTCGA 3732delA synt F T(15)GGAGATGACATCTGGCCCTCAGGGGG 3732delA synt R T(15)CTCCTTCACGTGTGAATTCTCAATAA 3791delC synt F T(15)AAGAAGGTGGAAATGCCATATTAGAG 3791delC synt R T(15)TTGTATTTTGCTGTGAGATCTTTGAC 3821delT synt F T(15)ATTCCTTCTCAATAAGTCCTGGCCAG 3821delT synt R T(15)GAATGTTCTCTAATATGGCATTTCCA Q1238X synt F T(15)TAGAGGGTGAGATTTGAACACTGCTT Q1238X synt R T(15)AGCCAGGACTTATTGAGAAGGAAATG S1255X (ex19)synt F T(15)GTCTGGCCCTCAGGGGGCCAAATGAC S1255X (ex19) synt R T(15)CGTCATCTTTCTTCACGTGTGAATTC S1255X;L synt F T(15)AAGCTTTTTTGAGACTACTGAACACT S1255X;L synt R T(15)TATAACAAAGTAATCTTCCCTGATCC 3849 ϩ 4AϾG synt F T(15)GGATTTGAACACTGCTTGCTTTGTTA 3849 ϩ 4AϾG synt R T(15)CCACCCTCTGGCCAGGACTTATTGAG 3850 - 1GϾA synt F T(15)AGTGGGCCTCTTGGGAAGAACTGGAT 3850 - 1GϾA synt R T(15)TTATAAGGTAAAAGTGATGGGATCAC 3905insT synt F T(15)TTTTTTTGAGACTACTGAACACTGAA 3905insT synt R T(15)AAAAAAAGCTGATAACAAAGTACTCT 3876delA synt F T(15)CGGGAAGAGTACTTTGTTATCAGCTT 3876delA synt R T(15)CGATCCAGTTCTTCCCAAGAGGCCCA G1244V synt F T(15)TAAGAACTGGATCAGGGAAGAGTACT G1244V synt R T(15)ACCAAGAGGCCCACCTATAAGGTAAA G1249E synt F T(15)AGAAGAGTACTTTGTTATCAGCTTTT G1249E synt R T(15)TCTGATCCAGTTCTTCCCAAGAGGCC S1251N synt F T(15)ATACTTTGTTATCAGCTTTTTTGAGACTACTG S1251N synt R T(15)TTCTTCCCTGATCCAGTTCTTCCCAA S1252P synt F T(15)CCTTTGTTATCAGCTTTTTTGAGACT S1252P synt R T(15)GACTCTTCCCTGATCCAGTTCTTCCC D1270N synt F T(15)AATGGTGTGTCTTGGGATTCAATAAC D1270N synt R T(15)TGATCTGGATTTCTCCTTCAGTGTTC W1282R synt F T(15)CGGAGGAAAGCCTTTGGAGTGATACC W1282R synt R T(15)GCTGTTGCAAAGTTATTGAATCCCAA R1283K synt F T(15)AGAAAGCCTTTGGAGTGATACCACAG R1283K synt R T(15)TTCCACTGTTGCAAAGTTATTGAATC 4005 ϩ 1GϾA synt F T(15)ATGAGCAAAAGGACTTAGCCAGAAAA 4005 ϩ 1GϾA synt R T(15)TCTGTGGTATCACTCCAAAGGCTTTC 4010del4 synt F T(15)GTATTTTTTCTGGAACATTTAGAAAAAACTTGG 4010del4 synt R T(15)AAAATACTTTCTATAGCAAAAAAGAAAAGAAGAA 4016insT synt F T(15)TTTTTTTCTGGAACATTTAGAAAAAACTTGG 4016insT synt R T(15)AAAAAAATAAATACTTTCTATAGCAAAAAAGAAAAGAAGA CFTRdele21 synt F T(15)TAGGTAAGGCTGCTAACTGAAATGAT CFTRdele21 synt R T(15)CCTATAGCAAAAAAGAAAAGAAGAAGAAAGTATG 4382delA synt F T(15)GAGAGAACAAAGTGCGGCAGTACGAT 4382delA synt R T(15)CTCTATGACCTATGGAAATGGCTGTT Bold, mutation allele of interest; bold and italicized, modified nucleotide. Login to comment

[hide] Spectrum of CFTR mutations in cystic fibrosis and ... Hum Mutat. 2000;16(2):143-56. Claustres M, Guittard C, Bozon D, Chevalier F, Verlingue C, Ferec C, Girodon E, Cazeneuve C, Bienvenu T, Lalau G, Dumur V, Feldmann D, Bieth E, Blayau M, Clavel C, Creveaux I, Malinge MC, Monnier N, Malzac P, Mittre H, Chomel JC, Bonnefont JP, Iron A, Chery M, Georges MD
Spectrum of CFTR mutations in cystic fibrosis and in congenital absence of the vas deferens in France.
Hum Mutat. 2000;16(2):143-56., [PMID:10923036]

Abstract [show]
We have collated the results of cystic fibrosis (CF) mutation analysis conducted in 19 laboratories in France. We have analyzed 7, 420 CF alleles, demonstrating a total of 310 different mutations including 24 not reported previously, accounting for 93.56% of CF genes. The most common were F508del (67.18%; range 61-80), G542X (2.86%; range 1-6.7%), N1303K (2.10%; range 0.75-4.6%), and 1717-1G>A (1.31%; range 0-2.8%). Only 11 mutations had relative frequencies >0. 4%, 140 mutations were found on a small number of CF alleles (from 29 to two), and 154 were unique. These data show a clear geographical and/or ethnic variation in the distribution of the most common CF mutations. This spectrum of CF mutations, the largest ever reported in one country, has generated 481 different genotypes. We also investigated a cohort of 800 French men with congenital bilateral absence of the vas deferens (CBAVD) and identified a total of 137 different CFTR mutations. Screening for the most common CF defects in addition to assessment for IVS8-5T allowed us to detect two mutations in 47.63% and one in 24.63% of CBAVD patients. In a subset of 327 CBAVD men who were more extensively investigated through the scanning of coding/flanking sequences, 516 of 654 (78. 90%) alleles were identified, with 15.90% and 70.95% of patients carrying one or two mutations, respectively, and only 13.15% without any detectable CFTR abnormality. The distribution of genotypes, classified according to the expected effect of their mutations on CFTR protein, clearly differed between both populations. CF patients had two severe mutations (87.77%) or one severe and one mild/variable mutation (11.33%), whereas CBAVD men had either a severe and a mild/variable (87.89%) or two mild/variable (11.57%) mutations.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
104 c 4016insT, G1244E, R1158X, 3120+1G>A, 1677delTA, I1234V, E831X, 5T, Q220X, E92K, G91R. Login to comment

[hide] Calnexin family members as modulators of genetic d... Semin Cell Dev Biol. 1999 Oct;10(5):473-80. Chevet E, Jakob CA, Thomas DY, Bergeron JJ
Calnexin family members as modulators of genetic diseases.
Semin Cell Dev Biol. 1999 Oct;10(5):473-80., [PMID:10597630]

Abstract [show]
The endoplasmic reticulum (ER) is an intracellular compartment devoted to the synthesis, segregation and folding of soluble and membrane secretory proteins. Some mutations in these proteins lead to their incorrect or incomplete folding in the ER. The ER has a quality control system which detects misfolded proteins and then specifies their fate. Some mutated proteins are retained in the ER wherein they accumulate (Russell bodies for misfolded immunoglobulin heavy chains, the PiZZ for alpha 1-antitrypsin), others are retrotranslocated from the ER and degraded by the cytosolic proteasomal system, and yet other proteins are eventually secreted (in AZC-treated cells). In this review we summarize the role of ER resident proteins in quality control of mutated secretory proteins.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
85 Potential involvement of calnexin and calreticulin in genetically-inherited diseases Protein Mutation Disease Calnexin Calreticulin Degradation ␣1-antitrypsin null Hong-Kong Emphysema q ref. 54 ᎐ q ref. 34 Ž .Liver disease Z-allele Emphysema q ref. 2 ᎐ q ref. 2 Ž .ref. 2 Lungrliver disease CFTR ⌬F508 ref. 51 Cystic fibrosis q ref. 51 ᎐ ᎐ G85E ref. 55 ND ND G91R ref. 55 ND ND Myeloperoxidase Y173C ref. 56 Decreased q ref. 57 q ref. 57 q ref. 56 W569R ref. 56 microbicidal response Factor VIII R2307Q ref. 58 Haemophilia q ref. 37 q ref. 37 Thyroglobulin Congenital q ref. 59 q ref. 59 ᎐ hyperthyroid goiter Figure 4. Login to comment
119 ref. 2 Lungrliver disease CFTR DF508 ref. 51 Cystic fibrosis q ref. 51 ] ] G85E ref. 55 ND ND G91R ref. 55 ND ND Myeloperoxidase Y173C ref. 56 Decreased q ref. 57 q ref. 57 q ref. 56 W569R ref. 56 microbicidal response Factor VIII R2307Q ref. 58 Haemophilia q ref. 37 q ref. 37 Thyroglobulin Congenital q ref. 59 q ref. 59 ] hyperthyroid goiter Figure 4. Login to comment

[hide] Cystic fibrosis transmembrane conductance regulato... Biophys J. 1998 Mar;74(3):1320-32. Mansoura MK, Smith SS, Choi AD, Richards NW, Strong TV, Drumm ML, Collins FS, Dawson DC
Cystic fibrosis transmembrane conductance regulator (CFTR) anion binding as a probe of the pore.
Biophys J. 1998 Mar;74(3):1320-32., [PMID:9512029]

Abstract [show]
We compared the effects of mutations in transmembrane segments (TMs) TM1, TM5, and TM6 on the conduction and activation properties of the cystic fibrosis transmembrane conductance regulator (CFTR) to determine which functional property was most sensitive to mutations and, thereby, to develop a criterion for measuring the importance of a particular residue or TM for anion conduction or activation. Anion substitution studies provided strong evidence for the binding of permeant anions in the pore. Anion binding was highly sensitive to point mutations in TM5 and TM6. Permeability ratios, in contrast, were relatively unaffected by the same mutations, so that anion binding emerged as the conduction property most sensitive to structural changes in CFTR. The relative insensitivity of permeability ratios to CFTR mutations was in accord with the notion that anion-water interactions are important determinants of permeability selectivity. By the criterion of anion binding, TM5 and TM6 were judged to be likely to contribute to the structure of the anion-selective pore, whereas TM1 was judged to be less important. Mutations in TM5 and TM6 also dramatically reduced the sensitivity of CFTR to activation by 3-isobutyl 1-methyl xanthine (IBMX), as expected if these TMs are intimately involved in the physical process that opens and closes the channel.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
27 Glycine 91 was one of three residues in TM1 identified as being accessible to water-soluble thiol reagents, and this residue is also the site of a patient mutation, G91R, associated with a pancreatic sufficient phenotype (Guillermit et al., 1993). Login to comment
62 Expression levels Wild-type and 11 mutant CFTR constructs were used in this study: G91A, G91E, G91R, G314A, G314D, G314E, G314Q, K335R, K335A, K335D, and K335E. Login to comment
137 Permeability Ratios Wild type 4-9 3.42 Ϯ 0.28 1.42 Ϯ 0.04 1.22 Ϯ 0.02 0.39 Ϯ 0.01 0.44 Ϯ 0.03 G91A 3-6 3.24 Ϯ 0.26 1.53 Ϯ 0.04 1.27 Ϯ 0.02 0.37 Ϯ 0.04 0.40 Ϯ 0.04 G91E 3-7 3.50 Ϯ 0.54 1.59 Ϯ 0.04 1.27 Ϯ 0.01 0.35 Ϯ 0.01 0.51 Ϯ 0.04 G91R 3-4 5.26 ؎ 0.46* 1.60 Ϯ 0.03 1.40 ؎ 0.01* 0.32 Ϯ 0.04 0.64 ؎ 0.04* G314A 3-4 2.87 Ϯ 0.17 1.45 Ϯ 0.03 1.19 Ϯ 0.02 0.31 Ϯ 0.03 0.33 Ϯ 0.03 G314D 4 3.42 Ϯ 0.34 1.44 Ϯ 0.05 1.25 Ϯ 0.04 0.33 Ϯ 0.03 0.51 Ϯ 0.05 G314E 3-4 3.72 Ϯ 0.56 1.65 ؎ 0.09* 1.35 ؎ 0.03* 0.49 Ϯ 0.04 0.53 Ϯ 0.04 G314Q 3-4 3.89 Ϯ 0.37 1.62 Ϯ 0.11 1.27 Ϯ 0.04 0.36 Ϯ 0.03 0.62 Ϯ 0.05 K335R 3-5 3.44 Ϯ 0.29 1.35 Ϯ 0.04 1.22 Ϯ 0.03 0.40 Ϯ 0.05 0.41 Ϯ 0.07 K335A 5-6 5.34 ؎ 0.58* 1.48 Ϯ 0.06 1.28 Ϯ 0.04 0.37 Ϯ 0.03 0.60 Ϯ 0.06 K335D 4-6 3.02 Ϯ 0.19 1.50 Ϯ 0.03 1.10 ؎ 0.02* 0.54 ؎ 0.04* 0.65 ؎ 0.06* K335E 5-8 3.64 Ϯ 0.21 1.48 Ϯ 0.06 1.29 Ϯ 0.03 0.46 Ϯ 0.04 1.10 ؎ 0.04* B. Conductance Ratios Wild type 4-9 0.14 Ϯ 0.02 0.75 Ϯ 0.02 0.64 Ϯ 0.02 0.52 Ϯ 0.03 0.18 Ϯ 0.03 G91A 3-6 0.14 Ϯ 0.01 0.77 Ϯ 0.02 0.61 Ϯ 0.02 0.47 Ϯ 0.02 0.19 Ϯ 0.02 G91E 3-7 0.15 Ϯ 0.03 0.73 Ϯ 0.02 0.60 Ϯ 0.01 0.50 Ϯ 0.04 0.30 Ϯ 0.02 G91R 3-4 0.14 Ϯ 0.00 0.84 Ϯ 0.01 0.63 Ϯ 0.01 0.32 ؎ 0.01* 0.14 Ϯ 0.01 G314A 3-4 0.30 Ϯ 0.09 0.89 ؎ 0.01* 0.66 Ϯ 0.01 0.48 Ϯ 0.09 0.24 Ϯ 0.01 G314D 4 0.28 Ϯ 0.05 0.82 Ϯ 0.01 0.70 Ϯ 0.02 0.49 Ϯ 0.06 0.27 Ϯ 0.03 G314E 3-4 0.62 ؎ 0.07* 1.18 ؎ 0.04* 0.84 ؎ 0.05* 0.42 Ϯ 0.05 0.29 Ϯ 0.09 G314Q 3-4 0.63 ؎ 0.02* 1.01 ؎ 0.04* 0.82 ؎ 0.03* 0.50 Ϯ 0.02 0.42 ؎ 0.02* K335R 3-5 0.14 Ϯ 0.01 0.76 Ϯ 0.03 0.61 Ϯ 0.02 0.59 Ϯ 0.06 0.16 Ϯ 0.03 K335A 6 0.20 Ϯ 0.03 0.77 Ϯ 0.02 0.61 Ϯ 0.02 0.45 Ϯ 0.03 0.21 Ϯ 0.02 K335D 4-6 0.65 ؎ 0.04* 1.25 ؎ 0.02* 0.89 ؎ 0.02* 0.61 Ϯ 0.06 0.58 ؎ 0.06* K335E 5-8 0.50 ؎ 0.06* 1.19 ؎ 0.03* 0.89 ؎ 0.02* 0.53 Ϯ 0.03 0.48 ؎ 0.03* (A) The apparent permeability ratios (PS/PCl) for each substitute anion were calculated from the shift in reversal potential using the Goldman-Hodgkin-Katz relation (noted in Materials and Methods). Login to comment
150 Of the three substitutions for G91, the arginine (G91R) altered the RR most dramatically, increasing it nearly sevenfold, although the negatively charged glutamate (G91E) FIGURE 2 The effect of replacement of [Cl- ]o by [SCN- ]o is shown for wtCFTR (A) and G314E (B). Login to comment
163 Finally, the mutant that exhibited the most striking shape change (G91R) did not differ from wild type in its dose-dependent activation by IBMX (see below). Login to comment
165 Concentration-dependent activation of G91A, G91E, and G91R CFTR was not discernibly different from that of wtCFTR. Login to comment
169 TABLE 4 Quantitative analyses of the macroscopic I-V shape changes Mutant ⌬ Net charge n RR g(ϩ30)/g(-30) RR/RRWT Wild type 5 1.220 Ϯ 0.06 1.00 G91A 0 4 1.293 Ϯ 0.06 1.06 G91E -1 5 1.512 ؎ 0.10* 1.24 G91R 1 4 8.041 ؎ 0.87* 6.59 G314A 0 4 1.201 Ϯ 0.09 0.98 G314D -1 4 1.362 Ϯ 0.08 1.12 G314E -1 7 1.405 Ϯ 0.08 1.15 G314Q 0 5 1.376 Ϯ 0.10 1.13 K335R 0 4 1.209 Ϯ 0.06 0.99 K335A -1 4 1.295 Ϯ 0.07 1.06 K335D -2 5 0.762 ؎ 0.02* 0.62 K335E -2 4 0.919 ؎ 0.02* 0.75 The slope conductance was measured at ϩ30 mV and -30 mV with respect to the reversal potential. Login to comment
173 TABLE 5 Concentration-dependent activation of wtCFTR, G91, G314, and K335 variants by IBMX in the presence of 10 ␮M forskolin Mutant n K1/2(IBMX) (mM) Wild type 15 0.35 Ϯ 0.04 G91A 5 0.42 Ϯ 0.06 G91E 8 0.51 ؎ 0.06* G91R 5 0.49 Ϯ 0.09 G314A 10 1.21 ؎ 0.11* G314D 3 1.35 ؎ 0.16* G314E 8 6.39 ؎ 1.35* G314Q 4 14.26 ؎ 6.64* K335R 4 0.46 Ϯ 0.04 K335A 2 0.35 Ϯ 0.15 K335D 7 0.87 ؎ 0.13* K335E 3 0.95 ؎ 0.07* The steady-state slope conductance was measured at -60 mV as increasing concentrations of IBMX (0.02-5.0 mM) were added to the perfusate in the continued presence of 10 mM forskolin. Login to comment
198 The results presented here are consistent with the notion that the binding of anions within the CFTR pore is a sensitive indicator of changes in pore structure whereas permeability ratios appear to be rather insensitive to similar TABLE 6 Qualitative summary of the functional consequences of mutations at G91, G314, and K335 Property G91 (TM1) K335 (TM6) G314 (TM5) G91A G91E G91R K335R K335A K335D K335E G314A G314D G314E G314Q I-V shape - - ϩϩϩ - - ϩϩ ϩ - - - - Psub/PCl - - - - - - ϩϩ - - - - gsub/gCl - - - - - ϩϩϩ ϩϩϩ ϩϩ - ϩϩϩ ϩϩϩ SCN- binding - - - - - ϩϩϩ ϩϩϩ ϩϩ - ϩϩϩϩ ϩϩϩϩ Activation - - - - - ϩϩ ϩϩ ϩϩϩ ϩϩϩ ϩϩϩϩ ϩϩϩϩ Results are expressed as follows: -, function of the CFTR construct with the indicated substitution was indistinguishable from wild type; ϩ to ϩϩϩϩ, semiquantitative indication of the magnitude of the change in the function compared with wild type. Login to comment
262 One of the most interesting mutants surveyed in this study was the G91R CFTR, which exhibited a dramatic, nearly sevenfold, increase in outward rectification (Table 4) but was identical to wtCFTR with regard to the apparent binding of SCN and dose-dependent activation by IBMX. Login to comment
138 Permeability Ratios Wild type 4-9 3.42 afe; 0.28 1.42 afe; 0.04 1.22 afe; 0.02 0.39 afe; 0.01 0.44 afe; 0.03 G91A 3-6 3.24 afe; 0.26 1.53 afe; 0.04 1.27 afe; 0.02 0.37 afe; 0.04 0.40 afe; 0.04 G91E 3-7 3.50 afe; 0.54 1.59 afe; 0.04 1.27 afe; 0.01 0.35 afe; 0.01 0.51 afe; 0.04 G91R 3-4 5.26 d1e; 0.46* 1.60 afe; 0.03 1.40 d1e; 0.01* 0.32 afe; 0.04 0.64 d1e; 0.04* G314A 3-4 2.87 afe; 0.17 1.45 afe; 0.03 1.19 afe; 0.02 0.31 afe; 0.03 0.33 afe; 0.03 G314D 4 3.42 afe; 0.34 1.44 afe; 0.05 1.25 afe; 0.04 0.33 afe; 0.03 0.51 afe; 0.05 G314E 3-4 3.72 afe; 0.56 1.65 d1e; 0.09* 1.35 d1e; 0.03* 0.49 afe; 0.04 0.53 afe; 0.04 G314Q 3-4 3.89 afe; 0.37 1.62 afe; 0.11 1.27 afe; 0.04 0.36 afe; 0.03 0.62 afe; 0.05 K335R 3-5 3.44 afe; 0.29 1.35 afe; 0.04 1.22 afe; 0.03 0.40 afe; 0.05 0.41 afe; 0.07 K335A 5-6 5.34 d1e; 0.58* 1.48 afe; 0.06 1.28 afe; 0.04 0.37 afe; 0.03 0.60 afe; 0.06 K335D 4-6 3.02 afe; 0.19 1.50 afe; 0.03 1.10 d1e; 0.02* 0.54 d1e; 0.04* 0.65 d1e; 0.06* K335E 5-8 3.64 afe; 0.21 1.48 afe; 0.06 1.29 afe; 0.03 0.46 afe; 0.04 1.10 d1e; 0.04* B. Conductance Ratios Wild type 4-9 0.14 afe; 0.02 0.75 afe; 0.02 0.64 afe; 0.02 0.52 afe; 0.03 0.18 afe; 0.03 G91A 3-6 0.14 afe; 0.01 0.77 afe; 0.02 0.61 afe; 0.02 0.47 afe; 0.02 0.19 afe; 0.02 G91E 3-7 0.15 afe; 0.03 0.73 afe; 0.02 0.60 afe; 0.01 0.50 afe; 0.04 0.30 afe; 0.02 G91R 3-4 0.14 afe; 0.00 0.84 afe; 0.01 0.63 afe; 0.01 0.32 d1e; 0.01* 0.14 afe; 0.01 G314A 3-4 0.30 afe; 0.09 0.89 d1e; 0.01* 0.66 afe; 0.01 0.48 afe; 0.09 0.24 afe; 0.01 G314D 4 0.28 afe; 0.05 0.82 afe; 0.01 0.70 afe; 0.02 0.49 afe; 0.06 0.27 afe; 0.03 G314E 3-4 0.62 d1e; 0.07* 1.18 d1e; 0.04* 0.84 d1e; 0.05* 0.42 afe; 0.05 0.29 afe; 0.09 G314Q 3-4 0.63 d1e; 0.02* 1.01 d1e; 0.04* 0.82 d1e; 0.03* 0.50 afe; 0.02 0.42 d1e; 0.02* K335R 3-5 0.14 afe; 0.01 0.76 afe; 0.03 0.61 afe; 0.02 0.59 afe; 0.06 0.16 afe; 0.03 K335A 6 0.20 afe; 0.03 0.77 afe; 0.02 0.61 afe; 0.02 0.45 afe; 0.03 0.21 afe; 0.02 K335D 4-6 0.65 d1e; 0.04* 1.25 d1e; 0.02* 0.89 d1e; 0.02* 0.61 afe; 0.06 0.58 d1e; 0.06* K335E 5-8 0.50 d1e; 0.06* 1.19 d1e; 0.03* 0.89 d1e; 0.02* 0.53 afe; 0.03 0.48 d1e; 0.03* (A) The apparent permeability ratios (PS/PCl) for each substitute anion were calculated from the shift in reversal potential using the Goldman-Hodgkin-Katz relation (noted in Materials and Methods). Login to comment
151 Of the three substitutions for G91, the arginine (G91R) altered the RR most dramatically, increasing it nearly sevenfold, although the negatively charged glutamate (G91E) FIGURE 2 The effect of replacement of [Clafa; ]o by [SCNafa; ]o is shown for wtCFTR (A) and G314E (B). Login to comment

[hide] Co- and posttranslational translocation mechanisms... J Biol Chem. 1998 Jan 2;273(1):568-76. Lu Y, Xiong X, Helm A, Kimani K, Bragin A, Skach WR
Co- and posttranslational translocation mechanisms direct cystic fibrosis transmembrane conductance regulator N terminus transmembrane assembly.
J Biol Chem. 1998 Jan 2;273(1):568-76., [PMID:9417117]

Abstract [show]
Transmembrane topology of most eukaryotic polytopic proteins is established cotranslationally at the endoplasmic reticulum membrane through the action of alternating signal and stop transfer sequences. Here we demonstrate that the cystic fibrosis transmembrane conductance regulator (CFTR) achieves its N terminus topology through a variation of this mechanism that involves both co- and posttranslational translocation events. Using a series of defined chimeric and truncated proteins expressed in a reticulocyte lysate system, we have identified two topogenic determinants encoded within the first (TM1) and second (TM2) membrane-spanning segments of CFTR. Each sequence independently (i) directed endoplasmic reticulum targeting, (ii) translocated appropriate flanking residues, and (iii) achieved its proper membrane-spanning orientation. Signal sequence activity of TM1, however, was inefficient due to the presence of two charged residues, Glu92 and Lys95, located within its hydrophobic core. As a result, TM1 was able to direct correct topology for less than half of nascent CFTR chains. In contrast to TM1, TM2 signal sequence activity was both efficient and specific. Even in the absence of a functional TM1 signal sequence, TM2 was able to direct CFTR N terminus topology through a ribosome-dependent posttranslational mechanism. Mutating charged residues Glu92 and Lys95 to alanine improved TM1 signal sequence activity as well as the ability of TM1 to independently direct CFTR N terminus topology. Thus, a single functional signal sequence in either the first or second TM segment was sufficient for directing proper CFTR topology. These results identify two distinct and redundant translocation pathways for CFTR N terminus transmembrane assembly and support a model in which TM2 functions to ensure correct topology of CFTR chains that fail to translocate via TM1. This novel arrangement of topogenic information provides an alternative to conventional cotranslational pathways of polytopic protein biogenesis.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
43 Plasmid pSPCFTR(E92A/ K95A) was generated by PCR amplification of pSPCFTR(E92A) (sense primer (SP6 promoter) ATTTAGGTGACACTATAG, and antisense primer TACTGCAGCGGTGACGGCGCCTAA), digestion of the PCR fragment with AvaI/PstI (PstI encoded in antisense oligonucleotides) and ligation of the fragment into an AvaI/PstI digested pSPCFTRK95A vector. Plasmids pSPCFTR(G85E) and pSPCFTR(G91R) are described elsewhere (33). Login to comment
44 Plasmids TM1.P, TM1.P(G85E), TM1.P(G91R), TM1.P(E92A), TM1.P(E95A), and TM1.P(E92A/E95A) were constructed by PCR amplification of WT or corresponding mutant CFTR plasmids (sense primer (SP6 promoter), antisense primer TAGATAGGTCACCATAGAGCGTTCCTCCT) and ligation of HindIII/BstEII-digested PCR fragments into a HindIII/BstEII-digested vector, S.L.ST.gG.P (described in Ref. Login to comment
82 In contrast to WT chains, no protease-protected fragments were observed for TM1.P(G85E) or TM1.P(G91R) chains, demonstrating that these inherited cystic fibrosis-related mutations abolished the ability of TM1 to direct translocation of the P reporter (lanes 4-6 and 7-9, respectively). Login to comment
95 Plasmids TM1.P, TM1.P(G85E), TM1.P(G91R), TM1.P(E92A), TM1.P(K95A), and TM1.P- (E92A/K95A) were expressed in rabbit reticulocyte lysate supplemented with canine pancreas microsomal membranes (A) or in microinjected Xenopus oocytes (B) as described under "Materials and Methods." Login to comment
103 located); (ii) G85E and G91R mutations essentially abolished TM1 signal sequence activity (Ͻ5% of chains translocated); and (iii) E92A and E92A/K95A mutations improved TM1 signal sequence activity (36% and 70% of chains translocated, respectively). Login to comment
105 This, however, was not the case because greater than 70% of WT as well as G85E and G91R chains achieved their correct N terminus topology in the ER membrane ((33) and Fig. 4). Login to comment
144 Plasmids encoding TM1 mutations, G85E and G91R (33), were also included for comparison. Login to comment
191 Furthermore, even chains containing a completely defective TM1 signal sequence (G85E and G91R mutants) were properly oriented in the membrane but only if a functional TM2 signal sequence was present. Login to comment
45 Plasmids TM1.P, TM1.P(G85E), TM1.P(G91R), TM1.P(E92A), TM1.P(E95A), and TM1.P(E92A/E95A) were constructed by PCR amplification of WT or corresponding mutant CFTR plasmids (sense primer (SP6 promoter), antisense primer TAGATAGGTCACCATAGAGCGTTCCTCCT) and ligation of HindIII/BstEII-digested PCR fragments into a HindIII/BstEII-digested vector, S.L.ST.gG.P (described in Ref. 18). Login to comment
83 In contrast to WT chains, no protease-protected fragments were observed for TM1.P(G85E) or TM1.P(G91R) chains, demonstrating that these inherited cystic fibrosis-related mutations abolished the ability of TM1 to direct translocation of the P reporter (lanes 4-6 and 7-9, respectively). Login to comment
96 Plasmids TM1.P, TM1.P(G85E), TM1.P(G91R), TM1.P(E92A), TM1.P(K95A), and TM1.P- (E92A/K95A) were expressed in rabbit reticulocyte lysate supplemented with canine pancreas microsomal membranes (A) or in microinjected Xenopus oocytes (B) as described under "Materials and Methods." Login to comment
104 located); (ii) G85E and G91R mutations essentially abolished TM1 signal sequence activity (,5% of chains translocated); and (iii) E92A and E92A/K95A mutations improved TM1 signal sequence activity (36% and 70% of chains translocated, respectively). Login to comment
106 This, however, was not the case because greater than 70% of WT as well as G85E and G91R chains achieved their correct N terminus topology in the ER membrane ((33) and Fig. 4). Login to comment

[hide] Association of domains within the cystic fibrosis ... Biochemistry. 1997 Feb 11;36(6):1287-94. Ostedgaard LS, Rich DP, DeBerg LG, Welsh MJ
Association of domains within the cystic fibrosis transmembrane conductance regulator.
Biochemistry. 1997 Feb 11;36(6):1287-94., [PMID:9063876]

Abstract [show]
The cystic fibrosis transmembrane conductance regulator (CFTR) is a Cl- channel composed of two membrane-spanning domains (MSD), two nucleotide-binding domains (NBD), and an R domain. To understand how these domains interact, we expressed various constructs of CFTR containing membrane-spanning and/or cytosolic domains either separately or together. We then tested for functional association of these domains using the SPQ halide-efflux assay or physical association using coimmunoprecipitation experiments. Coexpression of the amino-terminal half (MSD1, NBD1, and the R domain) and the carboxy-terminal half (MSD2 and NBD2) of CFTR generated functional Cl- channel activity whereas expression of either alone did not give a signal with the SPQ assay. This result suggests that the two halves associate in the membrane. Using domain-specific antibodies, we found that either half of CFTR could coimmunoprecipitate the other, suggesting a physical association. Coimmunoprecipitation persisted between halves missing the NBDs, the R domain, or the amino-terminal tail. Moreover, constructs from MSD1 containing only the first and second transmembrane sequences and intervening extracellular loop were sufficient for interaction with MSD2. These data suggest that interactions between the two membrane-spanning domains of CFTR may mediate association between the two halves of the protein.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
207 We made an N-terminal construct that contained the CF-associated mutation G91R (Guillermit et al., 1993) in M1 (1-835G91R) and expressed it alone or with the C-terminal half, 837-1480. Login to comment

[hide] SSCP analysis: a blind sensitivity trial. Hum Mutat. 1997;10(1):65-70. Jordanova A, Kalaydjieva L, Savov A, Claustres M, Schwarz M, Estivill X, Angelicheva D, Haworth A, Casals T, Kremensky I
SSCP analysis: a blind sensitivity trial.
Hum Mutat. 1997;10(1):65-70., [PMID:9222762]

Abstract [show]
Studies of the sensitivity of SSCP analysis usually have been performed under conditions contrary to the rules of quality control trials and have produced widely different results. We have performed a blind trial of the sensitivity of SSCP analysis for the detection of mutations in fragments up to 500 bp in length under a fixed single set of electrophoretic conditions. The mutation detection rate was 84%. In addition, we have identified a second mutation in nine samples. All these mutations are polymorphisms, including a novel polymorphism 1248 + 52T/C first reported in the present work.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
58 SSCP pattern of different probes, screened for exon 3 of the CFTR gene. Line 1: control sample of known mutation (G91R), which is difficult to detect by SSCP analysis. Lines 2-14: original codes of samples, which according to the referral laboratory do not carry mutation in the exon under investigation. Login to comment

[hide] Haplotype analysis of 94 cystic fibrosis mutations... Hum Mutat. 1996;8(2):149-59. Morral N, Dork T, Llevadot R, Dziadek V, Mercier B, Ferec C, Costes B, Girodon E, Zielenski J, Tsui LC, Tummler B, Estivill X
Haplotype analysis of 94 cystic fibrosis mutations with seven polymorphic CFTR DNA markers.
Hum Mutat. 1996;8(2):149-59., [PMID:8844213]

Abstract [show]
We have analyzed 416 normal and 467 chromosomes carrying 94 different cystic fibrosis (CF) mutations with polymorphic genetic markers J44, IVS6aGATT, IVS8CA, T854, IVS17BTA, IVS17BCA, and TUB20. The number of mutations found with each haplotype is proportional to its frequency among normal chromosomes, suggesting that there is no preferential haplotype in which mutations arise and thus excluding possible selection for specific haplotypes. While many common mutations in the worldwide CF population showed absence of haplotype variation, indicating their recent origins, some mutations were associated with more than one haplotype. The most common CF mutations, delta F508, G542X, and N1303K, showed the highest number of slippage events at microsatellites, suggesting that they are the most ancient CF mutations. Recurrence was probably the case for 9 CF mutations (R117H, H199Y, R347YH, R347P, L558S, 2184insA, 3272-26A-->G, R1162X, and 3849 + 10kbC-->T). This analysis of 94 CF mutations should facilitate mutation screening and provides useful data for studies on population genetics of CF.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
105 CFTR Haplotypes for Diallelic and Multiallelic DNA Markers for 94 CF Mutations" J44-GATT- 8CA-17BTA- No. of T854-TUB20 17BCA Mutation chromosomes % Normal Laboratory Reference 2-7-1-2 17-47-13 (55.4%) 17-46-13 17-45-13 17-34-13 17-32-13 17-31-14 17-31-13 17-29-14 17-28-13 16-48-13 16-46-14 16-46-13 16-45-13 16-44-13 16-35-13 16-33-13 16-32-13 16-31-14 16-31-13 16-30-13 16-29-13 16-26-13 16-25-13 16-24-13 14-31-13 1-7-2-1 17-7-17 (16.8%) R334W R334W 3860ins31 G1244E R1162X R1162X R1162X G91R MllOlK R347P R334W R117C E92K 3849+lOkbC+T 3293delA 1811+1.6kb A-tG 1811+1.6kb A-tG 2184insA P205S 3659delC G673X 11005R I336K W58S R347P W846X 405+1-A G178R 3905insT R1162X R347H 3100insA E60X 1078delT 4005+1-A K710X 1677delTA H199Y 3601-2AjG 3850-3T+G 3272-26A-tG 3850-1-A 1812-1-A R117H L1059X S492F Y1092X Y569H 3732delA C866Y 711+1G+T 711+1-T G85E 1949del84 2789+5-A H1085R W1282X R1066C 2043delG V456F 2 1 1 1 2 1 6 2 2 1 2 1 1 2 1 1 4 1 1 1 3 2 1 1 1 1 1 1 2 7 1 1 1 1 2 1 1 3 19 3 3 1 1 2 1 1 5 1 1 1 1 3 6 3 5 1 13 2 1 1 - 0.48 0.48 - - - 0.24 - - - 2.65 2.40 1.93 2.65 1.68 2.65 0.72 13.94 13.46 1.93 - 0.72 0.24 3.37 - b b fP fP fP t b,fb.fP h fb t h t h h fP fP b.h b h h b h h h h h fb fb,fP.t fP fP fP9t fP b t fPh b h fb b.fb,h fb*fP b,fP h h t h fb fb,fp,h.t fP fP fb t b.fP,t b,fb,h,t b f b h h fb b,fb.fP,h fP h h Gasparini et al. (1991b) Chilldn et al. (1993a) Devoto et al. (1991) Gasparini et al. (1991b) Dork et al. (1993a) Guillermit et al. (1993) Zielenski et al. (1993) Dean et al. (1990) Dork et al. (1994a) Nunes et al. (1993) Highsmith et al. (1994) Ghanem et al. (1994) Chilldn et al. (1995) Dork et al. (1994a) Dork et al. (1993a) Chilldn et al. (1993b) Kerem et al. (1990) Dork et al. (1994a) Dork et al. (1994a) Cuppenset al. (1993) Fanen et al. (1992) Maggio et al. (personal communication) Audrezet et al. (1993) Vidaud et al. (1990) Dork et al. (1993b) Zielenski et al. (1991a) Chilldn et al. (1994b) Malik et al. (personal communication) Cremonesi et at. Login to comment

[hide] Neonatal screening for cystic fibrosis: result of ... Hum Genet. 1995 Nov;96(5):542-8. Ferec C, Verlingue C, Parent P, Morin JF, Codet JP, Rault G, Dagorne M, Lemoigne A, Journel H, Roussey M, et al.
Neonatal screening for cystic fibrosis: result of a pilot study using both immunoreactive trypsinogen and cystic fibrosis gene mutation analyses.
Hum Genet. 1995 Nov;96(5):542-8., [PMID:8530001]

Abstract [show]
We have evaluated a two-tier neonatal cystic fibrosis (CF) screening of immunoreactive trypsinogen (IRT) followed by CFTR gene mutation analysis using a systematic scanning of exons 7, 10, and 11, and, if necessary, by direct DNA sequencing. Over an 18-month period we screened 32,300 neonates born in the western part of Britanny. The first tier, involving IRT screening at 3 days of age, utilizes a low elevation of the trypsinogen level (600 ng/ml), which is highly sensitive. The second tier, which corresponds to the exhaustive screening for mutations in three exons of the gene, is highly specific for this population (Britanny). The false positive rate is very low, and no false negatives have been reported to date. This strategy has allowed the identification of five novel alleles (V322A, V317A, 1806 del A, R553G, G544S).

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
80 Identification of novel mutations The systematic screening of exons 7, 10, and I I performed on each positive Guthrie card during this period has led us to identify five new mutations in the CFTR 30 545 % of non AF508 mutations 20 9 1717-1G->A 10 & & i i Esox G91R I 621+1G->T R117H 6b[ 7 905delG 1078 del T R347H 1221 det CT F311L R347L i10 i11i12 13 14a~l ! Login to comment

[hide] Screening Young syndrome patients for CFTR mutatio... Am J Respir Crit Care Med. 1995 Oct;152(4 Pt 1):1353-7. Friedman KJ, Teichtahl H, De Kretser DM, Temple-Smith P, Southwick GJ, Silverman LM, Highsmith WE Jr, Boucher RC, Knowles MR
Screening Young syndrome patients for CFTR mutations.
Am J Respir Crit Care Med. 1995 Oct;152(4 Pt 1):1353-7., [PMID:7551394]

Abstract [show]
Young syndrome is characterized by obstructive azoospermia associated with chronic sinobronchial disease of an infectious nature, but normal sweat-gland and pancreatic function as well as normal nasal potential differences. Congenital bilateral absence of the vas deferens (CBAVD) in some patients arises from mutations within the cystic fibrosis (CF) transmembrane regulator (CFTR) gene. Because of some similarities between Young syndrome, CF, and CBAVD, we evaluated 13 patients with Young syndrome, including screening for more than 30 different mutations within the CFTR gene. The mean age of the patients was 43 yr (range, 32 to 50 yr), and all were of northern European extraction. The sweat chloride concentration was normal in all patients (mean = 29 mEq/L; range, 8 to 43 mEq/L). Most had intermittent bronchial and sinus infections, but none was chronically colonized with Staphylococcus aureus or Pseudomonas aeruginosa. The FEV1 was normal or only mildly reduced in most patients (mean = 74%; range, 48 to 100% predicted). Of 26 Young syndrome chromosomes, we identified one with the recognized CF mutation delta F508. The incidence of CFTR mutations (1 in 26) did not differ significantly from the expected carrier frequency in this population. In summary, it is unlikely that the typical Young syndrome patient has a clinical disease associated with CFTR mutation on both alleles.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
78 Of the 13 Young syndrome patients, we identified one (Patient 5) who was het- CBAVD Dl152H D1270N G576A* R75Q* P67L Rl17H 3849 + 10 KB C > T G551S Rl17H Pancreatic Sufficient, Moderate Pulmonary Symptoms, Normal Sweat Chloride Concentrations Pancreatic Sufficient, Moderate Pulmonary Symptoms R347P 2789 + 5 G > A R334W G85E R347H R347L Rl17H G91R A455E S945L Y563N Q1291H R297Q R352Q L1065P 3850-3 T > G F1286S 3849 + 10 KB C > T TABLE 1 CFTR MUTATION SCREENING PANEL Severe M508 G551D R553X N1303K W1282X G542X 1717-1 G > A ~1507 R560T 3659deiC 621 + 1 G > T S549N TABLE 2 CLINICAL FEATURES OF YOUNG SYNDROME PATIENTS Patient Age Sweat CI- FEV, Paranasal Sputum No. Login to comment

[hide] Mutation analysis in 600 French cystic fibrosis pa... J Med Genet. 1994 Jul;31(7):541-4. Chevalier-Porst F, Bonardot AM, Gilly R, Chazalette JP, Mathieu M, Bozon D
Mutation analysis in 600 French cystic fibrosis patients.
J Med Genet. 1994 Jul;31(7):541-4., [PMID:7525963]

Abstract [show]
The cystic fibrosis transmembrane conductance regulator (CFTR) gene of 600 unrelated cystic fibrosis (CF) patients living in France (excluding Brittany) was screened for 105 different mutations. This analysis resulted in the identification of 86% of the CF alleles and complete genotyping of 76% of the patients. The most frequent mutations in this population after delta F508 (69% of the CF chromosomes) are G542X (3.3%), N1303K (1.8%), W1282X (1.5%), 1717-1G-->A (1.3%), 2184delA + 2183 A-->G (0.9%), and R553X (0.8%).

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
21 Among the 104 other CFTR mutations tested on the 373 non-AF508 CF chromosomes, none of the following 58 mutations were found: G91R, 435 insA, 444delA, D11OH, 556delA, 557delT, R297Q, 1154insTC, R347L, R352Q, Q359K/T360K, 1221delCT, G480C, Q493R, V520F, C524X, 1706dell7, S549R (A-C), S549N, S549I, G551S, 1784delG, Q552X, L558S, A559T, R560T, R560K, Y563N, P574H, 2307insA, 2522insC, 2556insAT, E827X, Q890X, Y913C, 2991de132 (Dork et al, personal communication), L967S, 3320ins5, 3359delCT, H1085R, R1158X, 3662delA, 3667del4, 3667ins4, 3732delA, 3737delA, W1204X, 3750delAG, I 1234V, Q1238X, 3850- 3T-+G, 3860ins31, S1255X, 3898insC, D1270N, R1283M, F1286S, 4005 + I G-A. Forty-six other mutations were found on at Distribution of CFTR mutations found in our sample ofpopulation (1200 CF chromosomes) Mutations tested No of CF chromosomes Haplotypes Method with the mutation XV2C-KM19 (% of total CF alleles) Exon 3: G85E 4 (033) 3C HinfI/ASO394delTT 2 2B PAGEExon 4: R117H 1 B ASOY122X 2 2C MseI/sequenceI148T 1 B ASO621+IG-J* 1 B MseIIASOExon 5: 711+1G--T 8(07) 8A ASOExon 7: AF311 1 C PAGE/sequencelO78delT 5 (0-42) 5C PAGE/ASOR334W 5 (0-42) 2A,2C,ID MspIlASOR347P 5 (042) 5A CfoI/NcoIR347H 1 Cfol/sequenceExon 9: A455E 1 B ASOExon 10: S492F I C DdeI/sequenceQ493X 1 D ASOl609deICA 1 C PAGE/Ddel/sequenceA1507 3 (025) 3D PAGE/ASOAF508 827 (69) 794B,30D,2C,IA PAGEl677delTA 1 A PAGE/sequenceExon I11: 1717-IG--.A 16(1-3) 14B Modified primers + AvaIIG542X 40 (3-3) 29B,5D,2A Modified primers + BstNiS549R(T--*G) 2 2B ASOG551D 3 (025) 3B HincII/Sau3AR553X 10(0-8) 6A,1B,2C,ID Hincll/sequenceExon 12: 1898+IG--A 1 C ASO1898+ IG-C 2 IC ASOExon 13: l9l8deIGC 1 A PAGE/sequence1949de184 I C PAGE/sequenceG628R(G-+A) 2 2A Sequence2118de14 I c PAGE/sequence2143de1T 1 B PAGE/modified primers2184de1A+2183A--*G 11 (0-9) lIB PAGE/ASO2184de1A 1 ASOK710X 3 (025) IC XmnI2372de18 1 B PAGE/sequenceExon 15: S945L 1 C TaqlExon 17b:L1065P I MnlIL1077P 1 A ASOY1092X 3 (025) 2C,IA Rsal/ASOExon 19: RI1162X 6 (0-5) 5C,IA DdeI/ASO3659delC 3 (025) 3C ASOExon 20: G1244E 2 2A MboIIS1251N 2 2C RsaI3905insT 4 (0-33) 4C PAGE/ASOW1282X 18 (105) 15B,1D MnlI/ASOR1283K 1 C Mnll/sequenceExon 21: N1303K 22 (1-8) 18B,lA,ID Modified primers+BstNI 47 mutations 1031 (85 9) least one CF chromosome (table): 21 of them are very rare as they were found on only one CF chromosome in our population. Login to comment

[hide] Amino acid residues lining the chloride channel of... J Biol Chem. 1994 May 27;269(21):14865-8. Akabas MH, Kaufmann C, Cook TA, Archdeacon P
Amino acid residues lining the chloride channel of the cystic fibrosis transmembrane conductance regulator.
J Biol Chem. 1994 May 27;269(21):14865-8., [PMID:7515047]

Abstract [show]
The cystic fibrosis transmembrane conductance regulator forms a chloride channel that is regulated by phosphorylation and intracellular ATP levels. The structure of the channel-forming domains is undetermined. To identify the residues lining this channel we substituted cysteine, one at a time, for 9 consecutive residues (91-99) in the M1 membrane-spanning segment. The cysteine substitution mutants were expressed in Xenopus oocytes. We determined the accessibility of the engineered cysteine to charged, sulfhydryl-specific methanethiosulfonate reagents added extracellularly. We assume that, among residues in membrane-spanning segments, only those lining the channel will be accessible to react with these hydrophilic reagents and that such a reaction would irreversibly alter conduction through the channel. Only the cysteines substituted for Gly-91, Lys-95, and Gln-98 were accessible to the reagents. We conclude that these residues are in the channel lining. The periodicity of these residues is consistent with an alpha-helical secondary structure.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
122 Although the Arg side chain is somewhat larger than theside chain of Cys modified by MTSEA', the reduction in whole cell current we observed following modification of G91C by MTSEA' suggests that the G91R mutant will have altered single-channel properties. Login to comment

[hide] A new missense mutation (G27E) in exon 2 of the CF... Hum Mol Genet. 1994 Feb;3(2):365-6. Bienvenu T, Cazeneuve C, Beldjord C, Dusser D, Kaplan JC, Hubert D
A new missense mutation (G27E) in exon 2 of the CFTR gene in a mildly affected cystic fibrosis patient.
Hum Mol Genet. 1994 Feb;3(2):365-6., [PMID:7516232]

Abstract [show]

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
35 This observation is consistent with the mild clinical phenotype already observed in compound heterozygote patients with another missense mutation in exon 3 (G91R) (13) or in homozygous patient for the G85E mutation (14). Login to comment

[hide] A novel mutation in exon 3 of the CFTR gene. Hum Genet. 1993 Apr;91(3):233-5. Guillermit H, Jehanne M, Quere I, Audrezet MP, Mercier B, Ferec C
A novel mutation in exon 3 of the CFTR gene.
Hum Genet. 1993 Apr;91(3):233-5., [PMID:7682984]

Abstract [show]
We have screened the 27 exons of the cystic fibrosis transmembrane conductance regulator gene in 87 non-delta F508 chromosomes of Breton origin using the combined techniques of denaturing gradient gel electrophoresis and direct sequencing. By this process, we have detected a new missense mutation, G91R, which results in an arginine for glycine at codon 91. Three affected patients with a delta F508/G91R genotype are pancreatic sufficient. Such observations could facilitate a better understanding of the functional importance of different regions of the encoded product and of the pathogenesis of the disease.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
2 By this process, we have detected a new missense mutation, G91R, which results in an arginine for glycine at codon 91. Login to comment
3 Three affected patients with a AF508/G91R genotype are pancreatic sufficient. Login to comment
37 G91R was found on two chromosomes among the 87 analysed and was associated with haplotype C in both cases (KMI9 = 1; XV2c = 2). Login to comment
39 Pattern of G91R mutation in denaturing gradient gel electrophoresis (gradient 0/50). Login to comment
42 Result of the sequencing of the exon 3 PCR product, the sequence of the mutated allele G91R, by the dideoxy method according to Sanger et al. (1977) displayed this genotype, as one of the above cases had an affected sibling. Login to comment
57 In this study, we have reported a new mutation located in exon 3 (G91R) of three patients, two of whom are siblings. Login to comment
59 All these patients carry the same genotype, are compound heterozygotes AF508/G91R and are pancreatic sufficient. Login to comment
65 Some point mutations located in the transmembrane domain, such as the G91R mutation that we have described in this paper, are good candidates for being associated with mild forms of the disease. Login to comment

[hide] Identification of 12 novel mutations in the CFTR g... Hum Mol Genet. 1993 Jan;2(1):51-4. Audrezet MP, Mercier B, Guillermit H, Quere I, Verlingue C, Rault G, Ferec C
Identification of 12 novel mutations in the CFTR gene.
Hum Mol Genet. 1993 Jan;2(1):51-4., [PMID:7683952]

Abstract [show]
Over 200 mutations, besides the deletion delta F508, have been identified in the CFTR gene and are known to cause CF. In order to characterize the molecular defects of non delta F508 CF chromosomes of various French origin, we have combined the techniques of denaturing gradient gel electrophoresis (DGGE) and direct sequencing to screen for mutations in the whole coding sequence of the CFTR gene corresponding to the 27 exons and their exon-intron boundaries. This approach enabled us to identify 12 novel mutations which are described here. We have systematically tested a large number of other nucleotide changes distributed in the 27 exons, each of them was clearly detected. These data support the notion that the DGGE conditions we have defined for screening coding sequence of the CFTR gene allows the identification of most of, if not all, the CFTR gene mutations.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
98 This notion has also been confirmed by a series of observations of the effects of missense mutations, such as Rl 17H (28), G91R (29) or R347L (this report), which are associated with pancreatic insufficiency and a milder form of the disease. Login to comment

[hide] Cystic fibrosis genotypes and views on screening a... Am J Hum Genet. 1992 Nov;51(5):943-50. Scriver CR, Fujiwara TM
Cystic fibrosis genotypes and views on screening are both heterogeneous and population related.
Am J Hum Genet. 1992 Nov;51(5):943-50., [PMID:1384327]

Abstract [show]

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
76 of CF chromosomes]) Distribution and Mutation (%) Askhenazi from Israel (Abeliovich et al. 1992 [94]; Shoshani et al. 1992 [95]): W1282X .......................................... AFS08 ............................................. G542X ............................................ 3849 + 10 kb, CT ............................ N1303K ........................................... Total ............................................ Celtic Bretons from France (Ferec et al. 1992 [365]): AF508 ............................................. 1078delT ......................................... G5S1D ............................................ 1717-1 G-A ..................................... W846X ............................................ G91R .............................................. Login to comment

[hide] The Transmission Interfaces Contribute Asymmetrica... J Biol Chem. 2015 Jul 3;290(27):16954-63. doi: 10.1074/jbc.M115.652602. Epub 2015 May 18. Loo TW, Clarke DM
The Transmission Interfaces Contribute Asymmetrically to the Assembly and Activity of Human P-glycoprotein.
J Biol Chem. 2015 Jul 3;290(27):16954-63. doi: 10.1074/jbc.M115.652602. Epub 2015 May 18., [PMID:25987565]

Abstract [show]
P-glycoprotein (P-gp; ABCB1) is an ABC drug pump that protects us from toxic compounds. It is clinically important because it confers multidrug resistance. The homologous halves of P-gp each contain a transmembrane (TM) domain (TMD) with 6 TM segments followed by a nucleotide-binding domain (NBD). The drug- and ATP-binding sites reside at the interface between the TMDs and NBDs, respectively. Each NBD is connected to the TMDs by a transmission interface involving a pair of intracellular loops (ICLs) that form ball-and-socket joints. P-gp is different from CFTR (ABCC7) in that deleting NBD2 causes misprocessing of only P-gp. Therefore, NBD2 might be critical for stabilizing ICLs 2 and 3 that form a tetrahelix bundle at the NBD2 interface. Here we report that the NBD1 and NBD2 transmission interfaces in P-gp are asymmetric. Point mutations to 25 of 60 ICL2/ICL3 residues at the NBD2 transmission interface severely reduced P-gp assembly while changes to the equivalent residues in ICL1/ICL4 at the NBD1 interface had little effect. The hydrophobic nature at the transmission interfaces was also different. Mutation of Phe-1086 or Tyr-1087 to arginine at the NBD2 socket blocked activity or assembly while the equivalent mutations at the NBD1 socket had only modest effects. The results suggest that the NBD transmission interfaces are asymmetric. In contrast to the ICL2/3-NBD2 interface, the ICL1/4-NBD1 transmission interface is more hydrophilic and insensitive to mutations. Therefore the ICL2/3-NBD2 transmission interface forms a precise hydrophobic connection that acts as a linchpin for assembly and trafficking of P-gp.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
315 The NBD2 transmission interface was also found to be particularly important for CFTR assembly as processing mutations in other domains that cause cystic fibrosis (such as èc;F508 in NBD1, G91R in TMD1, L1093P in TMD2) were found to impair the conformational stability of NBD2 (54). Login to comment

[hide] Development of CFTR Structure. Front Pharmacol. 2012 Sep 6;3:162. doi: 10.3389/fphar.2012.00162. eCollection 2012. Patrick AE, Thomas PJ
Development of CFTR Structure.
Front Pharmacol. 2012 Sep 6;3:162. doi: 10.3389/fphar.2012.00162. eCollection 2012., [PMID:22973227]

Abstract [show]
Cystic fibrosis is a lethal genetic disease caused by lack of functional cystic fibrosis transmembrane conductance regulator (CFTR) proteins at the apical surface of secretory epithelia. CFTR is a multidomain protein, containing five domains, and its functional structure is attained in a hierarchical folding process. Most CF-causing mutations in CFTR, including the most common mutation, a deletion of phenylalanine at position 508 (DeltaF508), are unable to properly fold into this functional native three dimensional structure. Currently, no high-resolution structural information about full length CFTR exists. However, insight has been gained through examining homologous ABC transporter structures, molecular modeling, and high-resolution structures of individual, isolated CFTR domains. Taken together, these studies indicate that the prevalent DeltaF508 mutation disrupts two essential steps during the development of the native structure: folding of the first nucleotide binding domain (NBD1) and its later association with the fourth intracellular loop (ICL4) in the second transmembrane domain (TMD2). Therapeutics to rescue DeltaF508 and other mutants in CFTR can be targeted to correct defects that occur during the complex folding process. This article reviews the structural relationships between CFTR and ABC transporters and current knowledge about how CFTR attains its structure-with a focus on how this process is altered by CF-causing mutations in a manner targetable by therapeutics.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
160 An example of mutants similarly located within CFTR with different local mechanisms of misfolding are the G85E and G91R mutations. Login to comment
165 The G91R mutant was predicted to have a similar effect on CFTR (Xiong et al., 1997), but this proved not to be true with regards to the TM1 conformation/integration profile (Patrick et al., 2011). Login to comment
166 Interestingly, the corrector compound four rescues G91R but not G85E-CFTR (Grove et al., 2009), suggesting the differences in the mutant molecular pathologies may be relevant for their ability to benefit from specific treatments to rescue defective CFTR. Login to comment

[hide] Mechanisms of CFTR Folding at the Endoplasmic Reti... Front Pharmacol. 2012 Dec 13;3:201. doi: 10.3389/fphar.2012.00201. eCollection 2012. Kim SJ, Skach WR
Mechanisms of CFTR Folding at the Endoplasmic Reticulum.
Front Pharmacol. 2012 Dec 13;3:201. doi: 10.3389/fphar.2012.00201. eCollection 2012., [PMID:23248597]

Abstract [show]
In the past decade much has been learned about how Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) folds and misfolds as the etiologic cause of cystic fibrosis (CF). CFTR folding is complex and hierarchical, takes place in multiple cellular compartments and physical environments, and involves several large networks of folding machineries. Insertion of transmembrane (TM) segments into the endoplasmic reticulum (ER) membrane and tertiary folding of cytosolic domains begin cotranslationally as the nascent polypeptide emerges from the ribosome, whereas posttranslational folding establishes critical domain-domain contacts needed to form a physiologically stable structure. Within the membrane, N- and C-terminal TM helices are sorted into bundles that project from the cytosol to form docking sites for nucleotide binding domains, NBD1 and NBD2, which in turn form a sandwich dimer for ATP binding. While tertiary folding is required for domain assembly, proper domain assembly also reciprocally affects folding of individual domains analogous to a jig-saw puzzle wherein the structure of each interlocking piece influences its neighbors. Superimposed on this process is an elaborate proteostatic network of cellular chaperones and folding machineries that facilitate the timing and coordination of specific folding steps in and across the ER membrane. While the details of this process require further refinement, we finally have a useful framework to understand key folding defect(s) caused by DeltaF508 that provides a molecular target(s) for the next generation of CFTR small molecule correctors aimed at the specific defect present in the majority of CF patients.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
66 An important implication of this topogenesis mechanism is highlighted by two CF-causing mutations, G85E and G91R, each of which introduces an additional ionizable residue into TM1. Login to comment
68 Despite achieving correct topology, however, G85E and G91R still disrupt CFTR folding and trafficking (Xiong et al., 1997; Patrick et al., 2011). Login to comment

[hide] Neonatal screening for cystic fibrosis: comparing ... J Cyst Fibros. 2014 Jul;13(4):384-90. doi: 10.1016/j.jcf.2014.01.004. Epub 2014 Feb 7. Sarles J, Giorgi R, Berthezene P, Munck A, Cheillan D, Dagorn JC, Roussey M
Neonatal screening for cystic fibrosis: comparing the performances of IRT/DNA and IRT/PAP.
J Cyst Fibros. 2014 Jul;13(4):384-90. doi: 10.1016/j.jcf.2014.01.004. Epub 2014 Feb 7., [PMID:24513262]

Abstract [show]
BACKGROUND: French health authorities promoted a study on 553,167 newborns comparing the performances of IRT/DNA and IRT/PAP for CF newborn screening. METHODS: In parallel to IRT/DNA, PAP was assayed in newborns with IRT>50 mug/L. Provisional PAP cutoffs at 3.0 mug/L when 50<IRT<100 mug/L and 1.7 mug/L when IRT>100 were used. Positive newborns were subjected to sweat test. Optimal cutoffs were established by a non-inferiority method. RESULTS: 95 CF newborns were identified (83 classical forms (ClF), including 9 meconium ileus (MI), and 12 atypical (mild) forms (AF) Of them, IRT/DNA identified 85 (73 ClF including 5 MI and 12 AF). PAP cutoffs at 1.8 mug/L when 50< IRT<100 mug/L and 0.6 mug/L when IRT>100 mug/L would identify 82 CF: 77 ClF, including 8 MI, and 5 AF. The number of sweat tests was 314 and 1039 in the IRT/DNA and IRT/PAP strategies, respectively. CONCLUSIONS: Using the optimal cutoffs, the sensitivity of the IRT/PAP strategy would not be inferior to that of IRT/DNA if identification of MF is not required.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
158 IRT d3 Ctrl IRT PAP Cl- Mut 1 Mut 2 1 66 68 0.4 80 ƊF508del ƊF508del 2 87.8 106.5 0.5 137 E1104X E1104X 3 93.2 105.8 0.8 82 G91R ƊF508del 4 71.1 56.7 0.3 80.0 ƊF508del ƊF508del 5 67.9 54.4 1.5 99.0 ƊF508del ƊF508del 6 87.1 82.9 4.5 70.0 E1104X D110H 7 61.5 62 5.0 88.0 R553X A455E 8 62.4 63.0 14.6 110.0 2183AANG 907delCins11 9 117.0 81.5 15.6 130.0 S466X S466X Lines 1-3: false negatives in the IRT/PAP strategy, 6-9: false negatives in the IRT/DNA strategy, due to mutations not detected by the Elucigeneࡊ CF30, 45: false negatives in both strategies. Login to comment

[hide] An image analysis method to quantify CFTR subcellu... Mol Cell Probes. 2014 Aug;28(4):175-80. doi: 10.1016/j.mcp.2014.02.004. Epub 2014 Feb 21. Pizzo L, Fariello MI, Lepanto P, Aguilar PS, Kierbel A
An image analysis method to quantify CFTR subcellular localization.
Mol Cell Probes. 2014 Aug;28(4):175-80. doi: 10.1016/j.mcp.2014.02.004. Epub 2014 Feb 21., [PMID:24561544]

Abstract [show]
Aberrant protein subcellular localization caused by mutation is a prominent feature of many human diseases. In Cystic Fibrosis (CF), a recessive lethal disorder that results from dysfunction of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR), the most common mutation is a deletion of phenylalanine-508 (pF508del). Such mutation produces a misfolded protein that fails to reach the cell surface. To date, over 1900 mutations have been identified in CFTR gene, but only a minority has been analyzed at the protein level. To establish if a particular CFTR variant alters its subcellular distribution, it is necessary to quantitatively determine protein localization in the appropriate cellular context. To date, most quantitative studies on CFTR localization have been based on immunoprecipitation and western blot. In this work, we developed and validated a confocal microscopy-image analysis method to quantitatively examine CFTR at the apical membrane of epithelial cells. Polarized MDCK cells transiently transfected with EGFP-CFTR constructs and stained for an apical marker were used. EGFP-CFTR fluorescence intensity in a region defined by the apical marker was normalized to EGFP-CFTR whole cell fluorescence intensity, rendering "apical CFTR ratio". We obtained an apical CFTR ratio of 0.67 +/- 0.05 for wtCFTR and 0.11 +/- 0.02 for pF508del. In addition, this image analysis method was able to discriminate intermediate phenotypes: partial rescue of the pF508del by incubation at 27 degrees C rendered an apical CFTR ratio value of 0.23 +/- 0.01. We concluded the method has a good sensitivity and accurately detects milder phenotypes. Improving axial resolution through deconvolution further increased the sensitivity of the system as rendered an apical CFTR ratio of 0.76 +/- 0.03 for wild type and 0.05 +/- 0.02 for pF508del. The presented procedure is faster and simpler when compared with other available methods and it is therefore suitable as a screening method to identify mutations that completely or mildly affect CFTR processing. Moreover, it could be extended to other studies on the biology underlying protein subcellular localization in health and disease.

Comments [show]

None has been submitted yet.

Sentences [show]
No. Sentence Comment
210 Structural cues involved in endoplasmic reticulum degradation of G85E and G91R mutant cystic fibrosis transmembrane conductance regulator. Login to comment