ABCMdb

ABCC7 p.Glu217Gly

ClinVar:	c.650A>G , p.Glu217Gly D , Pathogenic
CF databases:	c.650A>G , p.Glu217Gly (CFTR1) ? , The mutation was detected by heteroduplex analysis in a 2-year old male Polish patient with high sweat cloride (60-80 meq/l), pancreatic sufficiency, and moderate lung disease. His other CF mutation is unknown. It was also found by Yoshimura in 1999, in the CFTR alleles of a single patient with diffuse panbronchiolitis who has Q1352 H in the other allele.
Predicted by SNAP2:	A: N (66%), C: D (59%), D: N (93%), F: D (80%), G: N (66%), H: N (53%), I: D (59%), K: N (72%), L: D (59%), M: D (53%), N: N (82%), P: D (59%), Q: N (66%), R: D (66%), S: N (66%), T: N (66%), V: D (53%), W: D (85%), Y: D (75%),
Predicted by PROVEAN:	A: N, C: D, D: N, F: D, G: N, H: N, I: D, K: N, L: D, M: D, N: N, P: D, Q: N, R: N, S: N, T: N, V: D, W: D, Y: D,

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[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.

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474 Other mutations either affected the transport of LTC4 and/or E217G (Y1243F) or reduced resistance to drugs (Y1236F, T1241A and Y1243F), suggesting that residues of the cytoplasmic half of TM17 with side chain hydrogen bonding potential participate in the formation of a substrate-binding site [218]. Login to comment
469 Zhang et al. [234] have demonstrated that R1084 (TM14) caused a substantial reduction in E217G, LTC4 and GSH transport and drug-resistance but also in MTX and E2SO4 transport activity [233] but did not affect substrate binding. Login to comment
475 Mutation of W1246 (TM17) eliminated E217G transport and drug-resistance while LTC4 transport was not affected [228]. Login to comment
481 Zhang et al. [235] demonstrated that E1089 (TM14) was essential for anthracycline resistance and for transport conjugated organic anions such as LTC4 and E217G. Login to comment
482 T1242 (TM17) is important for E217G and LTC4 transport and a highly specific functional relationship between TM14 and TM17 for resistance to chemotherapeutic drugs. Login to comment
484 When mutated, another residue from CL7, P1150, reduced the transport of LTC4, GSH but enhanced MTX and E217G transport. Login to comment
499 Zelcer et al. [242] also reported that the transport of E217G, which binds to TM17 [220], was modulated by glibenclamide, probably via allosteric interaction. Login to comment

[hide] Distribution of CFTR gene mutations in cystic fibr... J Med Genet. 2000 Aug;37(8):E16. Teder M, Klaassen T, Oitmaa E, Kaasik K, Metspalu A
Distribution of CFTR gene mutations in cystic fibrosis patients from Estonia.
J Med Genet. 2000 Aug;37(8):E16., [PMID:10922396]

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26 The mixture was heated to 80°C, Table 1 Mutations identified in CF patients from Estonia Mutation Exon No of chromosomes Frequency (%) Method Reference F508 10 31 51.7 HA 6 394delTT 3 8 13.3 HA 7 359insT 3 1 SSCP 16 3659delC 19 1 SSCP 17 E217G 6a 1 DGGE 18 H117C 4 1 SSCP 19 I1005R 17a 1 SSCP 19 R1066H 17b 1 DGGE 20 S1196X 19 1 DGGE 21 S1235R 19 2 DGGE 22 Unidentified 12 Total 60 Table 2 Genotypes of the 30 CF patients from Estonia No of patients Genotype 8 F508/ F508 6 F508/394delTT 1 F508/I1005R 1 F508/359insT 1 F508/3659delC 1 F508/H117C 1 F508/R1066H 1 F508/S1196X 1 394delTT/394delTT 2 S1235R/U* 3 F508/U 1 E217G/U 3 U/U *Unidentified mutation Electronic letter of 4 www.jmedgenet.com cooled slowly to 30°C, and then divided into four sets of 60 µl. Login to comment

[hide] Disease-associated mutations in the extracytoplasm... J Biol Chem. 2001 May 4;276(18):14848-54. Epub 2001 Feb 6. Hammerle MM, Aleksandrov AA, Riordan JR
Disease-associated mutations in the extracytoplasmic loops of cystic fibrosis transmembrane conductance regulator do not impede biosynthetic processing but impair chloride channel stability.
J Biol Chem. 2001 May 4;276(18):14848-54. Epub 2001 Feb 6., 2001-05-04 [PMID:11278813]

Abstract [show]
Consistent with its function as a chloride channel regulated entirely from the cytoplasmic side of the plasma membrane, the cystic fibrosis transmembrane conductance regulator (CFTR) glycoprotein exposes little of its mass on the exterior surface of cells. The first and fourth extracytoplasmic loops (ELs) contain approximately 15 and 30 residues, respectively; the other four ELs are extremely short. To examine the influence of missense mutants in ELs detected in patients with cystic fibrosis, we have expressed them in mammalian (baby hamster kidney (BHK21)) cells and assessed their biosynthetic processing and chloride channel activity. In contrast to previous findings that 18 of 30 disease-associated missense mutations in cytoplasmic loops caused retention of the nascent polypeptides in the endoplasmic reticulum, all the EL mutants studied matured and were transported to the cell surface. This pronounced asymmetry is consistent with the notion that endoplasmic reticulum quality control of nascent CFTR is exerted primarily on the cytoplasmic side of the membrane. Although this set of EL mutations has little effect on CFTR maturation, most of them seriously compromise its chloride channel activity. Substitutions at six different positions in EL1 and single positions in EL2 and EL4 all destabilized the open state, some of them severely, indicating that the ELs contribute to the stability of the CFTR ion pore.

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75 TABLE I Oligonucleotide primers used to generate mutations Mutation Primer S108F GGAAGAATCATAGCTTtCTATGACCCGGATAAC Y109C AGAATCATAGCTTCCTgTGACCCGGATAACAAG D110H ATCATAGCTTCCTATcACCCGGATAACAAGGAG P111A ATAGCTTCCTATGACgCGGATAACAAGGAGGAA P111L ATAGCTTCCTATGACCtGGATAACAAGGAGGAA E116K CCGGATAACAAGGAGaAACGCTCTATCGCGATT R117C GATAACAAGGAGGAAtGCTCTATCGCGATTTAT R117H GATAACAAGGAGGAACaCTCTATCGCGATTTAT R117L GATAACAAGGAGGAACtCTCTATCGCGATTTAT R117P GATAACAAGGAGGAACcCTCTATCGCGATTTAT E217G ATGGGGCTAATCTGGGgGTTGTTACAGGCGTCT T908N TATGCAGTGATTATCAaCAGCACCAGTTCGTAT P1013L GTCGCAGTTTTACAACtCTACATCTTTGTTGCA FIG. 2. Login to comment
99 For example, E217G in EL2 reduces the efflux rate greatly. Login to comment
120 D, squares, E217G; circles, T908N; triangles, P1013L. Login to comment
152 Strikingly, the single missense mutation (E217G) in the short EL2 also results in channels with only transient rather than stable openings, perhaps implying that this loop is involved also in stabilizing open pore structure. Login to comment
171 For example a nucleotide binding domain mutation, G551D, precludes virtually all TABLE II Relative charge transport capacity of mutants Mutants S108F Y109C D110H P111L P111A E116K R117H R117C R117L R117P E217G T908N P1013L Imutant/Iwt 100% 11 15 27 173 105 12 80 27 5 11 10 48 170 FIG. 5. Login to comment
188 The E217G mutant in EL2 had a similar effect. Login to comment

[hide] Analysis of exocrine pancreatic function in cystic... Eur J Clin Invest. 2001 Sep;31(9):796-801. Walkowiak J, Herzig KH, Witt M, Pogorzelski A, Piotrowski R, Barra E, Sobczynska-Tomaszewska A, Trawinska-Bartnicka M, Strzykala K, Cichy W, Sands D, Rutkiewicz E, Krawczynski M
Analysis of exocrine pancreatic function in cystic fibrosis: one mild CFTR mutation does not exclude pancreatic insufficiency.
Eur J Clin Invest. 2001 Sep;31(9):796-801., [PMID:11589722]

Abstract [show]
BACKGROUND: Cystic fibrosis (CF) is the most common cause of exocrine pancreatic insufficiency in childhood. The aim of the present study is to evaluate the correlation between genotype and exocrine pancreatic insufficiency in CF patients. The special emphasis was put on the analysis of mild CFTR mutations. DESIGN: The study comprised 394 CF patients and 105 healthy subjects (HS). Elastase-1 concentrations were measured in all subjects. RESULTS: Severe pancreatic insufficiency was associated with the presence of two CFTR gene mutations (DeltaF508, N1303K, CFTR dele 2,3 (21kb), G542X, 1717-1G-A, R533X, W1282X, 621GT, 2183AAG, R560T, 2184insA and DeltaI507, G551D, 895T) and mild insufficiency with the presence of at least one mutation (R117H, 3171insC, A155P2, 138insL, 296 + 1G-A, E92GK, E217G, 2789 + 5G-A. 3849 + 1kbC-T/3849 + 1kbC-T) genotype resulted in high elastase-1-values. However, in case of patients with genotype DeltaF508/3849 + 10kbC-T, 1717-1GA/3849 + 10kbC-T as well as with DeltaF508/R334W, both high and low elastase-1 concentrations were found. Low E1 values were found in a patient with DeltaF508/R347P genotype. CONCLUSION: Patients who carry two 'severe' mutations develop pancreatic insufficiency, whereas those who carry at least one 'mild' usually remain pancreatic sufficient. However, the presence of one mild mutation does not exclude pancreatic insufficiency.

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5 Results Severe pancreatic insufficiency was associated with the presence of two CFTR gene mutations (DF508, N1303K, CFTR dele 2,3 (21kb), G542X, 1717±1G-A, R533X, W1282X, 621GT, 2183AAG, R560T, 2184insA and DI507, G551D, 895T) and mild insufficiency with the presence of at least one mutation (R117H, 3171insC, A155P2, 138insL, 296 1 1G-A, E92GK, E217G, 2789 1 5G-A. Login to comment
51 Results Among 394 genotyped CF patients, the following mutations on alleles were found (n): DF508 (464), 3849 1 10kbC-T (30), CFTR dele2,3(21 kB) (21), N1303K (15), G542X (12), 1717±1G-A (9), R533X (6), W1282X (6), 621 1 G-T (3), R117H (2), 3171insC (2), A155P2 (2), 2183AAG (2), R334W (2), 895T (2), 296 1 1G-A (2), E92GK (2), 138insL (1), E217G (1), 2789 1 5G-A (1), R347P (1), R560T (1), 2184insA (1), I507 (1), G551D (1). Login to comment
58 156 mg g21 ) ± mildly affecting pancreatic function (R117H, 3171insC, A155P2, 296 1 1G-A, E92GK, 138insL, E217G, 2789 1 5G-A). Login to comment
81 500 DF508/3849 1 10kbC-T (17) 1 4 1 6 5 DF508/CFTR dele2,3(21kb) (15) 9 4 2 DF508/N1303K (10) 7 3 DF508/1717±1G-A (7) 5 2 DF508/G542X (7) 4 2 1 DF508/W1282X (5) 4 1 DF508/R553X (3) 3 DF508/R334W (2) 1 1 DF508/2183AAG (2) 2 DF508/R117H (1) 1 DF508/621GT (1) 1 DF508/R347P (1) 1 DF508/2184insA (1) 1 DF508/DI507 (1) 1 3849 1 10kbC-T/3849 1 10kbC-T (3) 3 N1303K/CFTR dele2,3(21kb) (2) 1 1 1717±1G-A/3849 1 10kbC-T (2) 1 1 3171insC/A155P2 (2) 1 1 296 1 1G-A/E92GK (2) 2 R117H/138insL (1) 1 W1282X/3849 1 10kbC-T (1) 1 N1303K/3849 1 10kbC-T (1) 1 CFTR dele2,3(21kb)/3849 1 10kbC-T (1) 1 R553X/G542X (1) 1 621 1 1G-T/621 1 1G-T (1) 1 G542X/M (4) 2 2 CFTR dele 2,3(21kb)/M (1) 1 2 3849 1 10kbC-T/M (2) 1 1 R533X/M (2) 2 N1303K/M (2) 2 895T/M (2) 1 1 E217G/M (1) 1 G551D/M (1) 1 R560T/M (1) 1 2789 1 5G-A/M (1) 1 Total (109) 44 21 10 4 12 18 M, unidentified mutation. Login to comment
91 Based on our findings we are able to classify additional mutations as `severe' (CFTR dele2,3(21kb), 2183AAG, 895T, 2184insA) and `mild' (3171insC, A155P2, 138insL, 296 1 1G-A, E92GK, E217G, 2789 1 5G-A). Login to comment

[hide] Determination of the relative contribution of thre... Eur J Hum Genet. 2002 Feb;10(2):100-6. Audrezet MP, Chen JM, Le Marechal C, Ruszniewski P, Robaszkiewicz M, Raguenes O, Quere I, Scotet V, Ferec C
Determination of the relative contribution of three genes-the cystic fibrosis transmembrane conductance regulator gene, the cationic trypsinogen gene, and the pancreatic secretory trypsin inhibitor gene-to the etiology of idiopathic chronic pancreatitis.
Eur J Hum Genet. 2002 Feb;10(2):100-6., [PMID:11938439]

Abstract [show]
In the last 5 years, mutations in three genes, the cystic fibrosis transmembrane conductance regulator (CFTR) gene, the cationic trypsinogen (PRSS1) gene, and the pancreatic secretory trypsin inhibitor (PSTI) gene, have been found to be associated with chronic pancreatitis (CP). In this study, using established mutation screening methods, we systematically analysed the entire coding sequences and all exon/intron junctions of the three genes in 39 patients with idiopathic CP (ICP), with a view to evaluating the relative contribution of each gene to the aetiology of the disease. Our results demonstrate that, firstly, 'gain-of-function' mutations in the PRSS1 gene may occasionally be found in an obvious ICP subject. Secondly, presumably 'loss-of-function' mutations in the PSTI gene appear to be frequent, with a detection rate of at least 10% in ICP and, finally, abnormal CFTR alleles are common: at least 20% of patients carried one of the most common CFTR mutations, and about 10% of patients were compound heterozygotes, having at least one 'mild' allele. Thus, in total, about 30% of ICP patients carried at least one abnormal allele in one of the three genes, and this is the most conservative estimate. Moreover, a trans-heterozygous state with sequence variations in the PSTI/CFTR genes was found in three patients. However, an association between the 5T allele in intron 8 of the CFTR gene and ICP remains unproven.

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56 `Gain-of-function' PRSS1 mutations are rare in ICP While PRSS1 mutations are often found in patients with hereditary pancreatitis, they can also be identified in subjects with ICP, albeit with an exceptionally low Table 1 Sequence variations identified in the PRSS1, PSTI, and CFTR genes in 39 patients with ICP CFTR Patient PRSS1 PSTI Mutant PolyT 1 ± a ± ± 7T/7T 2 ± ± F508del/R352Q 9T/7T 3 ± ± F508del/P5L 9T/7T 4 ± ± c.4575+2G4A 9T/7T 5 ± ± ± 7T/7T 6 ± N34Sb ± 7T/7T 7 ± ± ± 7T/5T 8 ± ± F508del/Q1476X 9T/7T 9 ± ± ± 7T/7T 10 ± ± ± 7T/7T 11 ± ± ± 7T/7T 12 ± ± ± 7T/7T 13 ± ± V562I 7T/5T 14 ± ± 2C4A W1282X 7T/5T 15 ± ± IVS3-6T4C 7T/7T 16 R122H ± ± 7T/7T 17 ± ± ± 9T/7T 18 ± ± ± 7T/5T 19 ± ± ± 7T/7T 20 ± N34S/N34S ± 7T/7T 21 ± ± ± 9T/5T 22 ± ± ± 7T/7T 23 ± ± E217G/A1136T 9T/7T 24 ± ± ± 7T/7T 25 ± ± ± NDc 26 ± ± ± ND 27 ± N34S IVS18 ± 20T4C 9T/7T 28 ± ± F508del 9T/7T 29 ± ± ± 7T/7T 30 ± ± N1303K ND 31 ± ± G542X 9T/7T 32 ± ± ± 7T/5T 33 ± ± F508del 9T/7T 34 ± ± 41G4Ad ± 7T/7T 35 ± ± ± 9T/7T 36 ± ± ± 9T/7T 37 ± ± ± 7T/7T 38 ± N34S L967S 7T/7T 39 ± ± ± 7T/5T a Indicates two wild alleles. Login to comment
85 Firstly, we found a total of 10 additional alleles (R352Q, P5L, c.4575+2G4A, V562I, IVS3-6T4C, E217G/A1136T, IVS18-20T4C, and L967S; Table 1) that would have been missed by the conventional genotyping method. Login to comment

[hide] Cystic fibrosis: a worldwide analysis of CFTR muta... Hum Mutat. 2002 Jun;19(6):575-606. Bobadilla JL, Macek M Jr, Fine JP, Farrell PM
Cystic fibrosis: a worldwide analysis of CFTR mutations--correlation with incidence data and application to screening.
Hum Mutat. 2002 Jun;19(6):575-606., [PMID:12007216]

Abstract [show]
Although there have been numerous reports from around the world of mutations in the gene of chromosome 7 known as CFTR (cystic fibrosis transmembrane conductance regulator), little attention has been given to integrating these mutant alleles into a global understanding of the population molecular genetics associated with cystic fibrosis (CF). We determined the distribution of CFTR mutations in as many regions throughout the world as possible in an effort designed to: 1) increase our understanding of ancestry-genotype relationships, 2) compare mutational arrays with disease incidence, and 3) gain insight for decisions regarding screening program enhancement through CFTR multi-mutational analyses. Information on all mutations that have been published since the identification and cloning of the CFTR gene's most common allele, DeltaF508 (or F508del), was reviewed and integrated into a centralized database. The data were then sorted and regional CFTR arrays were determined using mutations that appeared in a given region with a frequency of 0.5% or greater. Final analyses were based on 72,431 CF chromosomes, using data compiled from over 100 original papers, and over 80 regions from around the world, including all nations where CF has been studied using analytical molecular genetics. Initial results confirmed wide mutational heterogeneity throughout the world; however, characterization of the most common mutations across most populations was possible. We also examined CF incidence, DeltaF508 frequency, and regional mutational heterogeneity in a subset of populations. Data for these analyses were filtered for reliability and methodological strength before being incorporated into the final analysis. Statistical assessment of these variables revealed that there is a significant positive correlation between DeltaF508 frequency and the CF incidence levels of regional populations. Regional analyses were also performed to search for trends in the distribution of CFTR mutations across migrant and related populations; this led to clarification of ancestry-genotype patterns that can be used to design CFTR multi-mutation panels for CF screening programs. From comprehensive assessment of these data, we offer recommendations that multiple CFTR alleles should eventually be included to increase the sensitivity of newborn screening programs employing two-tier testing with trypsinogen and DNA analysis.

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109 Mutational Arrays, Detection Rates and Methods by Region* Estimated Projected detection of Number of Number of Country/ allele two CFTR mutations chromosomes Region Mutation array detectiona mutationsb includedc (max/min)d Reference Europe Albania ∆F508 (72.4%) C276X (0.7%) 74.5 55.5 4 270/146 CFGAC [1994]; Macek et al. G85E (0.7%) R1070Q (0.7%) [2002] Austria ∆F508 (62.9%) 457TAT→G (1.2%) 76.6 58.7 11 1516/580 Estiville et al. [1997]; Dörk et al. (total) G542X (3.3%) 2183AA→G (0.7%) [2000]; Macek et al. [2002] CFTRdele2,3 (2.1%) N1303K (0.6%) R1162X (1.9%) I148T (0.5%) R553X (1.7%) R117H (0.5%) G551D (1.2%) Austria ∆F508 (74.6%) 2183AA→G (2.4%) 95.3 90.8 8 126 Stuhrmann et al. [1997] (tyrol) R1162X (8.7%) G551D (1.6%) G542X (2.4%) R347P (1.6%) 2789+5G→A (2.4%) Q39X (1.6%) Belarus ∆F508 (61.2%) R553X (0.5%) 75.2 56.6 9 278/188 Dörk et al. [2000]; Macek et al. G542X (4.5%) R334W (0.5%) [2002] CFTRdele2,3 (3.3%) R347P (0.5%) N1303K (3.2%) S549N (0.5%) W1282X (1.0%) Belgium ∆F508 (75.1%) 622-1A→C (0.5%) 100.0 100.0 27 1504/522 Cuppens et al. [1993]; Mercier et G542X (3.5%) G458V (0.5%) al. [1993]; CFGAC [1994]; N1303K (2.7%) 1898+G→C (0.5%) Estivill et al.[1997] R553X (1.7%) G970R (0.5%) 1717-1G→A (1.6%) 4218insT (0.5%) E60X (1.6%) 394delTT (0.5%) W1282X (1.4%) K830X (0.5%) 2183A→G+2184delA (1.2%) E822K (0.5%) W401X (1.0%) 3272-1G→A (0.5%) A455E (1.0%) S1161R (0.5%) 3272-26A→G (1.0%) R1162X (0.5%) S1251N (1.0%) 3750delAG (0.5%) S1235R (0.8%) S1255P (0.5%) ∆I507 (0.6%) Bulgaria ∆F508 (63.6%) R75Q (1.0%) 93.0 86.5 21 948/432 Angelicheva et al. [1997]; (total) N1303K (5.6%) 2183AA→G (0.9%) Estivill et al. [1997]; Macek G542X (3.9%) G1244V+S912L (0.9%) et al. [2002] R347P (2.2%) G85E (0.9%) 1677delTA (2.1%) 2184insA (0.9%) R1070Q (1.8%) L88X+G1069R (0.8%) Q220X (1.2%) 2789+5G→A (0.8%) 3849+10KbC→T (1.1%) G1244E (0.8%) W1282X (1.0%) 1717-1G→A (0.8%) 2176insC (1.0%) Y919C (0.7%) G1069R (1.0%) WORLDWIDEANALYSISOFCFTRMUTATIONS581 Bulgaria 1) DF508 4) 1677delTA - - 6 13 Angelicheva et al. [1997] (ethnic 2) R347P 5) Q493R Turks) 3) G542X 6) L571S - - 1 30 Angelicheva et al. [1997] Bulgaria 1) DF508 (100.0%) (Gypsy) Croatia ∆F508 (64.5%) G551D (1.1%) 72.5 52.6 5 276 Macek et al. [2002] G542X (3.3%) 3849+10KbC→T (0.7%) N1303K (2.9%) Czech ∆F508 (70.0%) 1898+1G→T (2.0%) 89.6 80.3 10 2196/628 CFGAC [1994]; Estiville et al. Republic CFTRdele2,3 (5.5%) 2143delT (1.2%) [1997]; Dörk et al. [2000]; G551D (3.8%) R347P (0.8%) Macek et al. [2002] N1303K (2.9%) 3849+10KbC→T (0.6%) G542X (2.2%) W1282X (0.6%) Denmark ∆F508 (87.5%) G542X (0.7%) 92.3 85.2 6 1888/678 CFGAC [1994]; Schwartz et al. (excluding 394delTT (1.8%) 621+1G→T (0.6%) [1994]; Estiville et al. [1997] Faroe) N1303K (1.1%) 3659delC (0.6%) Estonia ∆F508 (51.7%) R117C (1.7%) 80.2 64.3 10 165/80 Estivill et al. [1997]; Klaassen et 394delTT (13.3%) E217G (1.7%) al. [1998]; Macek et al. S1235R (3.3%) R1066H (1.7%) [2002] 359insT (1.7%) 3659delC (1.7%) I1005R (1.7%) S1169X (1.7%) Finland ∆F508 (46.2%) G542X (1.9%) 78.8 62.1 4 132/52 CFGAC [1994]; Kere et al. 394delTT (28.8%) 3372delA (1.9%) [1994]; Estivill et al. [1997] France ∆F508 (67.7%) 2789+5G→T (0.79%) 79.7 63.6 12 17854/7420 Chevalier-Porst et al. [1994]; (total) G542X (2.94%) 2184delA+2183A→G (0.77%) Estivill et al. [1997]; Claustres et al. [2000]; Guilloud-Bataille N1303K (1.83%) G551D (0.74%) et al. [2000] 1717-1G→A (1.35%) 1078delT (0.63%) W1282X (0.91%) ∆I507 (0.62%) R553X (0.86%) Y122K (0.59%) France ∆F508 (75.8%) R297Q (0.8%) 98.7 97.4 18 599/365 Férec et al. [1992]; Scotet et al. (Brittany) 1078delT (4.0%) R347H (0.8%) [2000] G551D (3.6%) I1234V (0.8%) N1303K (3.0%) R553X (0.8%) R117H (1.7%) 2789+5G→A (0.8%) 3272-26A→G (1.3%) 4005+1G→A (0.7%) G542X (1.1%) 621+1G→T (0.6%) 1717-1G→A (1.0%) ∆I507 (0.6%) G1249R (0.8%) W846X (0.5%) France ∆F508 (70.0%) N1303K (0.8%) 90.4 81.7 16 250 Claustres et al. [1993] (southern) G542X (6.4%) 3737delA (0.8%) 1717-1G→A (1.6%) R1162X (0.8%) L206W (1.2%) Y1092X (0.8%) R334W (1.2%) S945L (0.8%) ∆I507 (1.2%) K710X (0.8%) 2184delA (1.2%) 1078delT (0.8%) R1158X (1.2%) Y122X (0.8%) (Continued) BOBADILLAETAL. Login to comment

[hide] A haplotype-based molecular analysis of CFTR mutat... Hum Mol Genet. 2003 Sep 15;12(18):2321-32. Lee JH, Choi JH, Namkung W, Hanrahan JW, Chang J, Song SY, Park SW, Kim DS, Yoon JH, Suh Y, Jang IJ, Nam JH, Kim SJ, Cho MO, Lee JE, Kim KH, Lee MG
A haplotype-based molecular analysis of CFTR mutations associated with respiratory and pancreatic diseases.
Hum Mol Genet. 2003 Sep 15;12(18):2321-32., 2003-09-15 [PMID:12952861]

Abstract [show]
Aberrant membrane transport caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene is associated with a wide spectrum of respiratory and digestive diseases as well as cystic fibrosis. Using a gene scanning method, we found 11 polymorphisms and mutations of the CFTR gene in the Korean population. Individual variants at these sites were analyzed by conventional DNA screening in 117 control and 75 patients having bronchiectasis or chronic pancreatitis. In a haplotype determination based on a Bayesian algorithm, 15 haplotypes were assembled in the 192 individuals tested. Several haplotypes, especially with Q1352H, IVS8 T5, and E217G, were found to have disease associations in a case-control study. Notably, a common polymorphism of M470V appears to affect the intensity of the disease association. Among the two haplotypes having IVS8 T5, the T5-V470 haplotype showed higher disease association than the T5-M470 haplotype. In addition, a Q1352H mutation found in a V470 background showed the strongest disease association. The physiological significances of the identified mutations were rigorously analyzed. Non-synonymous E217G and Q1352H mutations in the M470 background caused a 60-80% reduction in CFTR-dependent Cl(-) currents and HCO3(-) -transport activities. Surprisingly, the additional M470V polymorphic variant with the Q1352H mutation completely abolished CFTR-dependent anion transport activities. These findings provide the first evidence on the importance of CFTR mutations in the Asian population. Importantly, the results also reveal that interactions between multiple genetic variants in cis affect the final function of the gene products.

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4 Several haplotypes, especially with Q1352H, IVS8 T5, and E217G, were found to have disease associations in a case-control study. Login to comment
9 Non-synonymous E217G and Q1352H mutations in the M470 background caused a 60-80% reduction in CFTR-dependent ClÀ currents and HCO3 À -transport activities. Login to comment
54 The frequency of E217G also seemed to be higher in bronchiectasis patients, especially in the subgroup of child bronchiectasis (P ¼ 0.04; ages under 18, three cases in 20 patients), although it was not significant when all patients were included (P ¼ 0.10; four cases in 47 patients). Login to comment
74 CFTR genetic variants analyzed in this study Variations found by TDGS Most common worldwide disease-causing mutations Reported disease-associated microsatellite À8G/C (50 UTR)a R117H (exon 4) T5-7,9 (IVS 8) (16) I125T (exon 4)b 621 þ 1G > T (intron 4) E217G (exon 6a)b F508del (exon 10) 1059C > T (exon 7, A309)a 1717-1G > A (intron 10) M470V (exon 10)b G542X (exon 11) I556V (exon 11)b G551D (exon 11) 2694T/G (exon 14a, T854)b R553X (exon 11) Q1352H (exon 22)b R1162X (exon 19) R1453W (exon 24)b W1282X (exon 20) N1303K (exon 21) Mutation names and nucleotide numbers are presented according to the Cystic Fibrosis Genetic Analysis Consortium (CFGAC; www.genet.sickkids.on.ca/). Login to comment
92 The frequency of haplotype 6 containing E217G was 5.8-fold higher in child bronchiectasis patients (7.5%, three in 40; P ¼ 0.03) than that in the control population (1.3%, three in 234), although this was not significant for all bronchiectasis patients (4.3%, four in 94; P ¼ 0.08). Login to comment
105 As shown in Figure 2A, the protein expressions of mature glycosylated CFTR were significantly decreased in E217G and Q1352H mutations. Login to comment
107 Frequency of CFTR gene variants in the Korean population Variation Genotype Group (number) Healthy control (n ¼ 117) Bronchiectasis (n ¼ 47) Pancreatitis (n ¼ 28) Diallelic -8G/C þ/þ 105 44 22 þ/Àa 12 3 6 R117H þ/þ 116 47 28 þ/À 1 0 0 I125T þ/þ 116 46 27 þ/À 1 1 1 E217G þ/þ 114 43 27 þ/À 3 4b 1 1059C > T þ/þ 117 47 27 (A309) þ/À 0 0 1 M470V þ/þ 23 3 6 þ/À 52 28 14 À/À 42 16 8 I556V þ/þ 111 45 28 þ/À 6 2 0 2694T/G þ/þ 41 16 8 (T854) þ/À 51 27 14 À/À 25 4 6 Q1352H þ/þ 116 43 24 þ/À 1 4* 4** R1453W þ/þ 115 46 28 þ/À 2 1 0 Microsatellite T5-7,9 5/7 4 6* 2 (IVS 8) 6/7 0 1 0 7/7 110 39*c 26 7/9 3 1 0 Differences between control and disease groups were analyzed by a chi-square test. When an expected cell value was less than 5, Fisher`s exact test was used. Login to comment
114 Haplotype assembly Allele ID -8G/C R117H I125T E217G 1059C/T T5-7,9 M470V I556V 2694T/G Q1352H R1453W Group a M470V-2694T/G background Control, n (%) Bronchiectasis, n (%) Pancreatitis, n (%) 1 G R I E C WTb V I T Q R 121 (51.7) 47 (50.0) 24 (42.9) 2-1 2 G R I E C WT M I G Q R 78 (33.3) 25 (26.6) 18 (32.1) 1-2 3 C R I E C WT M I G Q R 11 (4.7) 3 (3.2) 5 (8.9) 1-2 4 G R I E C WT V I T H R 1 (0.4) 4 (4.3)* 4 (7.1)** 2-1 5 G R I E C 5 V I T Q R 2 (0.9) 5 (5.4)* 1 (1.8) 2-1 6 G R I G C WT M I G Q R 3 (1.3) 4 (4.3)c 1 (1.8) 1-2 7 G R I E C WT V V T Q R 5 (2.1) 2 (2.2) 0 (0.0) 2-1 8 G R I E C WT V I G Q R 4 (1.7) 1 (1.0) 0 (0.0) 2-2 9 G R I E C 5 M I G Q R 2 (0.9) 1 (1.0) 1 (1.8) 1-2 10 G R I E C WT M I G Q W 2 (0.9) 1 (1.0) 0 (0.0) 1-2 11 G R T E C WT V I T Q R 1 (0.4) 1 (1.0) 1 (1.8) 2-1 12 G R I E C WT M I T Q R 2 (0.9) 0 (0.0) 0 (0.0) 1-1 13 C R I E C WT V I G Q R 1 (0.4) 0 (0.0) 0 (0.0) 2-2 14 G H I E C WT V V T Q R 1 (0.4) 0 (0.0) 0 (0.0) 2-1 15 C R I E T WT M I G Q R 0 (0.0) 0 (0.0) 1 (1.8) 1-2 Total 234 (100.0) 94 (100.0) 56 (100.0) Haplotypes were assembled using a software based on the Bayesian algorithm (Haplotyper) (7). Login to comment
122 Compared with WT, the expression levels of E217G and Q1352H were reduced by 64 and 73%, respectively, when CFTR band intensities were normalized against those of the cis-gene product DHFR (Fig. 2B). Login to comment
137 Therefore, it was concluded that the decreased current density in the whole cell ClÀ current of E217G (Fig. 3C) was due to the decreased membrane expression, and those of I556V and R1453W were due to the decreased Po. Login to comment
145 Compared to the WT, E217G and Q1352H showed significant reductions in forskolin-stimulated ClÀ /HCO3 À exchange by 65 and 77%, respectively. Login to comment
176 Characteristics of CFTR mutants selected for functional studies Name Nucleotide change Exon Domain Evolutionary conservationa Possible disease associationb E217G 782A >G 6a EC2 b, h, r CF with pancreatic sufficiency (Polish), Panbronchiolitis (Japanese) I556V 1798A >G 11 NBD1 All seven species Chronic bronchitis (French) Q1352H 4188G > C 22 NBD2 All seven species CBAVD (Japanese), Panbronchiolitis (Japanese) R1453W 4489C > T 24 IC10 b, h, m, r, s Panbronchiolitis (Japanese) a Evolutionary conservations are compared in CFTR genes of bovine (b), dogfish (d), human (h), mouse (m), rabbit (r), sheep (s), and xenopus (x). Login to comment
205 E217G was previously identified in a Polish CF patient with pancreatic sufficiency (data from CFGAC) and showed a marginal disease association in the present case-control study. Login to comment
206 These results imply that E217G is a mild mutation, which is also supported by our molecular data. Login to comment
207 The E217G mutation partially decreased membrane protein expression and anion transporting activities by 60-70%. Login to comment
208 However, the strongly disease associated Q1352H mutation also decreased the protein expression and functional activities by only 70-80%, which was not much different from E217G when they were measured in the same haplotype background of M470 (Figs 2-4). Login to comment
209 Of interest, E217G was found to arise in the haplotype having a high activity type of M470 (haplotype 6), and Q1352H to arise in the low activity type of V470 (haplotype 4) in the real population. Login to comment
226 In conclusion, CFTR mutations of M470V-Q1352H, IVS8 T5-M470V, and E217G are associated with bronchiectasis and chronic pancreatitis. Login to comment
259 The mutagenic primers were as follows: E217G, 50 -CTC CTC ATG GGG CTA ATC TGG GGG TTG TTA CAG GCG TCT G-30 M470V, 50 -CTG GAG CAG GCA AGA CTT CAC TTC TAA TGG TGA TTA TGG GAG-30 ; I556V, 50 -AGT GGA GGT CAA CGA GCA AGA GTT TCT TTA GCA AGG TGA AT-30 ; Q1352H, 50 -CCT AAG CCA TGG CCA CAA GCA CTT GAT GTG CTT GGC TAG-30 ; R1453W, 50 -GTG AAG CTC TTT CCC CAC TGG AAC TCA AGC AAG TGC AAG TCT-30 . Login to comment

[hide] The role of cystic fibrosis gene mutations in dete... Gastroenterol Clin North Am. 2004 Dec;33(4):817-37, vii. Cohn JA, Mitchell RM, Jowell PS
The role of cystic fibrosis gene mutations in determining susceptibility to chronic pancreatitis.
Gastroenterol Clin North Am. 2004 Dec;33(4):817-37, vii., [PMID:15528020]

Abstract [show]
This article reviews current concepts regarding the pathobiology of cystic fibrosis pancreatic disease. It summarizes recent studies on the relationship between CFTR mutations and pancreatitis, and it reviews several unresolved issues in the field.

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78 The European data Table 1 Abnormal CFTR and PSTI genotypes detected in two studies of idiopathic chronic pancreatitis* CFTR genotype category N Genotypes detected in individual subjects US study (Noone et al [47]) CFsev / CFm-v compound heterozygotes 8 DF508 / R117H-7T**; DF508 / 5T; DF508 / 5T; DF508 / D1152H; DF508 / D1152H; DF508 / P574H; DF508 / 3120G>A; 621þ1G>T/G1069R CFm-v / CFm-v compound heterozygotes 1 5T / 5T** CFsev / - (CF carriers) 1 N1303K / - CFm-v / - 7 R117H-7T / -; 5T / -**; 5T / -; 5T / -; 5T / -; 5T / -; 5T / - Normal (- / -) CFTR genotype 22 1 was homozygous for the N34S PSTI mutation; 5 were N34S carriers European study (Audrezet et al [50]) CFsev / CFm-v compound heterozygotes 4 DF508/R352Q; DF508/P5L; DF508/Q1476X; W1282X/5T*** CFm-v / CFm-v compound heterozygotes 2 V562I/5T; E217G/A1136T CFsev / - (CF carriers)**** 3 DF508 / -; DF508 / -; G542X / - CFm-v / - 9 L967S/-**; IVS18-20T>C/-**; c.4575þ2G>A/-; IVS3-6T>C; 5T/-; 5T/-; 5T/-; 5T/-; 5T/- Normal (- / -) CFTR genotype 17 1 was homozygous for the N34S PSTI mutation; 1 was a N34S carrier * CFTR mutations were classified as causing either severe (CFsev ) or mild-variable loss-of-function (CFm-v ) [18,47]; all detected CFsev mutations are CF-causing mutations according to current consensus criteria [79]. Login to comment

[hide] The impact of cystic fibrosis and PSTI/SPINK1 gene... Clin Lab Med. 2005 Mar;25(1):79-100. Cohn JA, Mitchell RM, Jowell PS
The impact of cystic fibrosis and PSTI/SPINK1 gene mutations on susceptibility to chronic pancreatitis.
Clin Lab Med. 2005 Mar;25(1):79-100., [PMID:15749233]

Abstract [show]
This article reviews current concepts regarding the pathobiology of cystic fibrosis pancreatic disease. It summarizes recent studies on the relationship between CFTR mutations and pancreatitis, and it reviews several unresolved issues in the field.

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90 Table 1 Abnormal CFTR and PSTI genotypes detected in two studies of idiopathic chronic pancreatitis* CFTR genotype category N Genotypes detected in individual subjects US study (Noone et al [47]) CFsev / CFm-v compound heterozygotes 8 DF508 / R117H-7T**; DF508 / 5T; DF508 / 5T; DF508 / D1152H; DF508 / D1152H; DF508 / P574H; DF508 / 3120G>A; 621þ1G>T/G1069R CFm-v / CFm-v compound heterozygotes 1 5T / 5T** CFsev / - (CF carriers) 1 N1303K / - CFm-v / - 7 R117H-7T / -; 5T / -**; 5T / -; 5T / -; 5T / -; 5T / -; 5T / - Normal (- / -) CFTR genotype 22 1 was homozygous for the N34S PSTI mutation; 5 were N34S carriers European study (Audrezet et al [50]) CFsev / CFm-v compound heterozygotes 4 DF508/R352Q; DF508/P5L; DF508/Q1476X; W1282X/5T*** CFm-v / CFm-v compound heterozygotes 2 V562I/5T; E217G/A1136T CFsev / - (CF carriers)**** 3 DF508 / -; DF508 / -; G542X / - CFm-v / - 9 L967S/-**; IVS18-20T>C/-**; c.4575þ2G>A/-; IVS3-6T>C; 5T/-; 5T/-; 5T/-; 5T/-; 5T/- Normal (- / -) CFTR genotype 17 1 was homozygous for the N34S PSTI mutation; 1 was a N34S carrier * CFTR mutations were classified as causing either severe (CFsev ) or mild-variable loss-of-function (CFm-v ) [18,47]; all detected CFsev mutations are CF-causing mutations according to current consensus criteria [79]. Login to comment

[hide] Reduced CFTR function and the pathobiology of idio... J Clin Gastroenterol. 2005 Apr;39(4 Suppl 2):S70-7. Cohn JA
Reduced CFTR function and the pathobiology of idiopathic pancreatitis.
J Clin Gastroenterol. 2005 Apr;39(4 Suppl 2):S70-7., [PMID:15758663]

Abstract [show]
Idiopathic chronic pancreatitis (ICP) is the leading cause of chronic pancreatitis in children and nonalcoholic adults. The risk of developing ICP is increased in individuals who have mutations of the cystic fibrosis gene (CFTR) and of a trypsin inhibitor gene (PSTI). In studies from the United States and France, the risk of ICP is increased about 40-fold by having two abnormal copies of the CFTR gene, about 14-fold by having the N34S PSTI mutation, and about 500-fold by having both. When ICP patients have two abnormal copies of the CFTR gene, there is also evidence of reduced residual CFTR protein function in extrapancreatic tissues based on clinical findings and nasal ion transport responses. Thus, pancreatitis risk is highest in individuals who have abnormalities in both the pancreatic ducts (CFTR) and acini (PSTI). These findings indicate that PSTI is a modifier gene for CFTR-related ICP and have implications for the diagnosis and pathogenesis of pancreatitis.

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69 Abnormal CFTR and PSTI Genotypes Detected in Two Studies of ICP CFTR Genotype Category* N Genotypes Detected in Individual Subjects U.S. study (Noone et al47 ) CFsev / CFm-v compound heterozygotes 8 DF508 / R117H-7T †; DF508 / 5T; DF508 / 5T; DF508 / D1152H; DF508 / D1152H; DF508 / P574H; DF508 / 3120G.A; 621 + 1G.T/G1069R CFm-v / CFm-v compound heterozygotes 1 5T / 5T † CFsev / 2 (CF carriers) 1 N1303K / 2 CFm-v / 2 7 R117H-7T / 2; 5T / 2 †; 5T / 2; 5T / 2; 5T / 2; 5T / 2; 5T / 2 Normal (2 / 2) CFTR genotype 22 1 was homozygous for the N34S PSTI mutation; 5 were N34S carriers French study (Audrezet et al50 ) CFsev / CFm-v compound heterozygotes 4 DF508/R352Q; DF508/P5L; DF508/Q1476X; W1282X/5T‡ CFm-v / CFm-v compound heterozygotes 2 V562I/5T; E217G/A1136T CFsev / 2 (CF carriers)§ 3 DF508 / 2; DF508 / 2; G542X / 2 CFm-v / 2 9 L967S/2 †; IVS18-20T.C/ 2†; c.4575+2G.A/2; IVS3-6T.C; 5T/2; 5 /2; 5T/ 2; 5T/2; 5T/ 2 Normal (2 / 2) CFTR genotype 17 1 was homozygous for the N34S PSTI mutation; 1 was a N34S carriers *Mutations of the cystic fibrosis (CF) gene (CFTR) were classified as causing either severe (CFsev ) or mild-variable loss-of-function (CFm-v )18,47 ; all detected CFsev mutations are CF-causing mutations according to current consensus criteria.68 In the U.S. study, most patients were tested for rare mutations by DNA sequencing47 ; in the French study, most patients were tested by dHPL.50 †These patients were also carriers for the N34S mutation of a trypsin inhibitor gene (PSTI). Login to comment

[hide] Gender-sensitive association of CFTR gene mutation... Mol Hum Reprod. 2005 Aug;11(8):607-14. Epub 2005 Aug 26. Morea A, Cameran M, Rebuffi AG, Marzenta D, Marangon O, Picci L, Zacchello F, Scarpa M
Gender-sensitive association of CFTR gene mutations and 5T allele emerging from a large survey on infertility.
Mol Hum Reprod. 2005 Aug;11(8):607-14. Epub 2005 Aug 26., [PMID:16126774]

Abstract [show]
Human infertility in relation to mutations affecting the cystic fibrosis transmembrane regulator (CFTR) gene has been investigated by different authors. The role of additional variants, such as the possible forms of the thymidine allele (5T, 7T and 9T) of the acceptor splice site of intron 8, has in some instances been considered. However, a large-scale analysis of the CFTR gene and number of thymidine residues, alone and in combination, in the two sexes had not yet been addressed. This was the aim of this study. Two groups were compared, a control group of 20,532 subjects being screened for perspective reproduction, and the patient group represented by 1854 idiopathically infertile cases. Analyses involved PCR-based CFTR mutations assessment, reverse dot-blot IVS8-T polymorphism analyses, denaturing gradient gel electrophoresis (DGGE) and DNA sequencing. The expected 5T increase in infertile men was predominantly owing to the 5/9 genotypic class. The intrinsic rate of 5T fluctuated only slightly among groups, but some gender-related differences arose when comparing their association. Infertile men showed a significantly enriched 5T + CFTR mutation co-presence, distributed in the 5/9 and 5/7 classes. In contrast, females, from both the control and the infertile groups, showed a trend towards a pronounced reduction of such association. The statistical significance of the difference between expected and observed double occurrence of 5T + CFTR traits in women suggests, in line with other reports in the literature, a possible survival-hampering effect. Moreover, regardless of the 5T status, CFTR mutations appear not to be involved in female infertility. These results underline the importance of (i) assessing large sample populations and (ii) considering separately the two genders, whose genotypically opposite correlations with these phenomena may otherwise tend to mask each other.

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76 This test involved nine subjects from the infertile group, revealing the occurrence of the following rare mutations: E217G, T1054A, W356X, D443Y and 3667insTC in males and L997F and R297Q in females and 29 subjects from the control, in which we found: A1009T, D110Y, E826K, G1069R, G1130A, G194V, I556V, L320F, M348K, M82V, P1290T, R117C, R352W, R74W, S42F, S660T, S911R, S912L, T1086A, T582S, V920L and Y89C. Login to comment

[hide] Identification of CFTR, PRSS1, and SPINK1 mutation... Pancreas. 2006 Oct;33(3):221-7. Keiles S, Kammesheidt A
Identification of CFTR, PRSS1, and SPINK1 mutations in 381 patients with pancreatitis.
Pancreas. 2006 Oct;33(3):221-7., [PMID:17003641]

Abstract [show]
OBJECTIVES: Chronic pancreatitis is a progressive inflammatory disorder leading to irreversible exocrine and/or endocrine impairment. It is well documented that mutations in the cationic trypsinogen (PRSS1) gene can cause hereditary pancreatitis. Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) and the serine protease inhibitor Kazal type 1 (SPINK1) genes are also associated with pancreatitis. METHODS: We analyzed 381 patients with a primary diagnosis of chronic or recurrent pancreatitis using the Ambry Test: Pancreatitis to obtain comprehensive genetic information for the CFTR, SPINK1, and PRSS1 genes. RESULTS: The results identified 32% (122/381) of patients with 166 mutant CFTR alleles, including 12 novel CFTR variants: 4375-20 A>G, F575Y, K598E, L1260P, G194R, F834L, S573C, 2789 + 17 C>T, 621+83 A>G, T164S, 621+25 A>G, and 3500-19 G>A. Of 122 patients with CFTR mutations, 5.5% (21/381) also carried a SPINK1 mutation, and 1.8% (7/381) carried a PRSS1 mutation. In addition, 8.9% (34/381) of all patients had 1 of 11 different SPINK1 mutations. Another 6.3% (24/381) of the patients had 1 of 8 different PRSS1 mutations. Moreover, 1.3% of the patients (5/381) had 1 PRSS1 and 1 SPINK1 mutation. A total 49% (185/381) of the patients carried one or more mutations. CONCLUSIONS: Comprehensive testing of the CFTR, PRSS1, and SPINK1 genes identified genetic variants in nearly half of all subjects considered by their physicians as candidates for genetic testing. Comprehensive test identified numerous novel variants that would not be identified by standard clinical screening panels.

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71 Patients With 1 CFTR Mutation CFTR Mutation 1 No. of Patients 1717-1 G9A 1 2789+5 G9A 1 3849+10kb C9T 2 3849+45 G9A 1 621+3 A9G 2 A1364V 1 A349V 1 A455E 1 D1152H 1 D1445N 1 deltaF508 16 E217G 1 F1286C 1 F316L 1 G542X 1 G551D 1 I148T 1 I807M 1 L206W 1 L967S 2 L997F 2 P55S 1 Q179K 1 Q220X 1 R117H 3 R1453W 1 R297Q 1 R31C 1 R668C 2 S1235R 1 S573C 1 S945L 1 V562A 1 V754M 2 Y1092X 1 Total patients 58 MutationsinboldfacewouldnothavebeendetectedbytheACOG/ACMGmutationpanel. Login to comment

[hide] Role of the extracellular loop in the folding of a... Biochemistry. 2007 Jun 19;46(24):7099-106. Epub 2007 May 22. Wehbi H, Rath A, Glibowicka M, Deber CM
Role of the extracellular loop in the folding of a CFTR transmembrane helical hairpin.
Biochemistry. 2007 Jun 19;46(24):7099-106. Epub 2007 May 22., 2007-06-19 [PMID:17516627]

Abstract [show]
The folding of membrane-spanning domains into their native functional forms depends on interactions between transmembrane (TM) helices joined by covalent loops. However, the importance of these covalent linker regions in mediating the strength of helix-helix associations has not been systematically addressed. Here we examine the potential structural impact of cystic fibrosis-phenotypic mutations in the extracellular loop 2 (ECL2) on interactions between the TM3 and TM4 helices of the cystic fibrosis transmembrane conductance regulator (CFTR) in constructs containing CFTR residues 194-241. When the effects of replacements in ECL2 (including the CF-phenotypic mutants E217G and Q220R) were evaluated in a library of wild-type and mutant TM3-ECL2-TM4 hairpin constructs, we found that SDS-PAGE gel migration rates differed over a range of nearly 40% +/- the wild-type position and that decreased migration rates correlate with increasing hairpin alpha-helical content as measured by circular dichroism spectra in sodium dodecyl sulfate micelles. The decreased mobility of TM3/4 constructs by introduction of non-native residues is interpreted in terms of an elongation or "opening" of the helical hairpin and concomitant destabilization of membrane-based helix-helix interactions. Our results support a role for short loop regions in dictating the stability of membrane protein folds and highlight the interplay between membrane-embedded helix-helix interactions and loop conformation in influencing the structure of membrane proteins.

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3 When the effects of replacements in ECL2 (including the CF-phenotypic mutants E217G and Q220R) were evaluated in a library of wild-type and mutant TM3-ECL2-TM4 hairpin constructs, we found that SDS-PAGE gel migration rates differed over a range of nearly 40% +/- the wild-type position and that decreased migration rates correlate with increasing hairpin R-helical content as measured by circular dichroism spectra in sodium dodecyl sulfate micelles. Login to comment
57 Two CF-phenotypic missense mutations, E217G and Q220R, have been documented in the ECL2 region (contributed to the Cystic Fibrosis Mutation Database at www.genet.sickkids.on.ca/cftr/ by Zielenski et al. and Fe´rec). Login to comment
59 E217G and Q220R point mutants were constructed initially in the TM3/4 WT background, and their migration on SDS-PAGE was compared to WT following the gel shift assay developed in our laboratory (33). Login to comment
61 We noted that the E217G and Q220R substitutions in the WT background each showed slower migration relative to WT on SDS-PAGE (Figure 2), initially implying that replacement of these ECL2 positions has rendered the TM3/4 WT construct less compact (vide infra). Login to comment
69 We therefore hypothesized that the S222/E217 pair might form an intraloop interaction that stabilizes the native hairpin conformation and systematically replaced each side chain with Gly in the double mutant cycle consisting of S222G, E217G, S222G/E217G, and E217S/S222E. Login to comment
70 The unequal migration rates of the S222G and E217G hairpins in this cycle (6.8 ( 0.7 vs 12.0 ( 2.1, p ) 0.015), however, were inconsistent with an interaction between the two residues, and the double mutant E217S/S222E, where the positions of the hydrogen bond donor and acceptor are interchanged, exhibited the largest perturbation in hairpin compactness (percent increase of apparent molecular weight compared to wild type ) 26%) than any others encountered in this work. Login to comment
78 (A) SDS-PAGE of TM3/4 WT and TM3/4 WT with E217G and with Q220R. Login to comment
97 When the changes in TM3/4 WT hairpin migration were compared to changes in overall hairpin helicity, a strong correlation (R ) 0.79) was observed (Figure 5), leading us to propose that increases in non-native R-helix structure within ECL2 might Table 1: Migration Behavior on SDS-PAGE Gels of Single and Double Mutants in the Loop Region of CFTR TM3/4 Constructs % change in apparent MW on SDS-PAGE mutant vs TM3/4 WT in WT loop mutantsa vs TM3/4 V232D in V232D loop mutantsa Pb E217G 6.8 ( 0.7 E217S 11.1 ( 3.4 5.4 ( 1.4 0.056 Q220R 15.2 ( 1.1 Q220G 0.3 ( 0.4 Q220N 2.1 ( 1.3 0.5 ( 0.3 0.108 Q220K 14.1 ( 1.0 Q220W 13.1 ( 1.3 11.5 ( 0.9 0.157 Q220E -11.1 ( 1.1 -4.0 ( 0.3 <0.001 S222G 12.0 ( 2.1 1.1 ( 0.6 0.001 S222E -0.3 ( 2.4 1.3 ( 0.5 0.512 E217G/S222G 12.4 ( 1.9 E217S/S222E 26.1 ( 4.5 averagec 10.4 ( 7.3 4.0 ( 4.2 0.067 a Values are the percentage difference vs TM3/4 WT or TM3/4 V232D migration of SDS-PAGE gels. Login to comment
147 For example, S222E and WT migrate at approximately the same rate, but Q220E moves at -11% vs WT; both S222G and E217G/S222G are at +12%; Q220K, Q220R, and Q220W are each at +13-15%. Login to comment
163 While conformational changes induced by ECL2 mutants in the TM3/4 hairpin should be minimized by additional intrachain contacts imposed by the full-length CFTR molecule, our results suggest that the molecular mechanism of disease introduced by the CF-phenotypic ECL2 mutants E217G and Q220R may involve introduction of non-native R-helical loop structure that destabilizes the CFTR fold and leads to aberrant function in the resting state and/or during substrate transport. Login to comment

[hide] Spectrum of mutations and variants/haplotypes of C... Clin Genet. 2007 Jun;71(6):530-9. Chang MC, Chang YT, Wei SC, Tien YW, Liang PC, Jan IS, Su YN, Wong JM
Spectrum of mutations and variants/haplotypes of CFTR and genotype-phenotype correlation in idiopathic chronic pancreatitis and controls in Chinese by complete analysis.
Clin Genet. 2007 Jun;71(6):530-9., [PMID:17539902]

Abstract [show]
Mutations in cystic fibrosis transmembrane conductance regulator (CFTR) gene have been reported in patients with chronic pancreatitis. The authors examine whether the mutations and haplotypes of CFTR will increase the risk of developing idiopathic chronic pancreatitis (ICP) in Chinese and their genotype and phenotype correlations. Seventy-eight patients with ICP and 200 geographically and ethnically matched controls in Taiwan were analyzed. The entire 27 coding and intronic regions of the CFTR gene were identified using heteroduplex analytical techniques and confirmed by sequencing analysis. The presence of 125G/C, 1001+10C>T, IVSTn(TG)m, 1540A>G, c2694T>G, and c4521G>A were determined by directing sequencing. Abnormal CFTR allele was found to be thrice as frequent in ICP patients as in controls (22/156 vs 19/400, p < 0.0001). T5 allele was associated with early onset of ICP. In six-loci haplotype analysis, 13 common haplotypes were assembled in the 278 individuals tested. The 125G/1001+11C/TG12/470M/2694T/4521G haplotype was associated with risk of ICP (odds ratio 11.3; 95% confidence interval 2.3-54.6, p = 0.008) in Chinese. The mutation spectrum is different from other ethnic groups. A population-specific panel of CFTR changes should be recommended for targeted populations including ICP in Chinese. It is important to design suitable screening programs for different populations.

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97 These mutations include I556V, G to A 3849145, N287Y, I125T, E217G, S895N, G1O69R, and Q1352H that have been found in patients with CP or CBAVD (http://www.genet. Login to comment
109 (%) in ICP Controls (%) I556V Exon 11 A to G 1798 Amino acid substitution 7 (8.9) 2 (1) IVS8-5T Intron 8 deletaion of 2T between 1342-12 and 1342-6 Aberrant splicing 6 (7.7) 14 (7) G to A 3849145 Intron 19 G to A at 3849145 mRNA splicing defect 3 (3.8) 2 (1) N287Y Exon 6b A to T 991 Amino acid substitution 2 (2.6) 00 (0) I125T Exon 4 T to C 506 Amino acid substitution 1 (1.3) 00 (0) E217G Exon 6a A to G 782 Amino acid substitution 1 (1.3) 00 (0) S895N Exon 15 G to A 2816 Missense mutation 1 (1.3) 00 (0) G1O69R Exon 17b G to A 3337 Amino acid substitution 1 (1.3) 00 (0) Q1352H Exon 22 G to C at 4188 Amino acid substitution 0 (0.0) 1 (0.5) ICP, idiopathic chronic pancreatitis. Login to comment
157 All our mutations are belonging to Ômild` mutations compatible with previous studies (6), including I556V, G to A 3849145, I125T, E217G, N287Y, S895N, G1O69R, and Q1352H. Login to comment
162 E217G was previously identified in a Polish CF patient with pancreatic sufficiency (data from CFGAC). Login to comment
163 The E217G mutation partially decreased membrane Table 4. Login to comment
170 Q1352H and E217G and M470V are considered to be strongly associated with chronic pancreatitis in Korean (9). Login to comment

[hide] Too much salt, too little soda: cystic fibrosis. Sheng Li Xue Bao. 2007 Aug 25;59(4):397-415. Quinton PM
Too much salt, too little soda: cystic fibrosis.
Sheng Li Xue Bao. 2007 Aug 25;59(4):397-415., 2007-08-25 [PMID:17700961]

Abstract [show]
Cystic fibrosis (CF) of the pancreas is the most widely accepted name of the most common fatal inherited single gene defect disease among Caucasians. Its incidence among other races is thought to be significantly less, but mutations in the gene have been reported in most, if not all, major populations. This review is intended to give general concepts of the molecular as well as physiological basis of the pathology that develops in the disease. First, an overview of the organ pathology and genetics is presented, followed by the molecular structure of the gene product (cystic fibrosis transmembrane conductance regulator, CFTR), its properties, functions, and controls as currently understood. Second, since mutations appear to be expressed primarily as a defect in electrolyte transport, effects and mechanisms of pathology are presented for two characteristically affected organs where the etiology is best described: the sweat gland, which excretes far too much NaCl ("salt") and the pancreas, which excretes far too little HCO3(- )("soda"). Unfortunately, morbidity and mortality in CF develop principally from refractory airway infections, the basis of which remains controversial. Consequently, we conclude by considering possible mechanisms by which defects in anion transport might predispose the CF lung to chronic infections.

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32 In surveys in Japan[31] , Korea[32] , and Vietnam[33] , no ΔF508 mutations were detected, but among Koreans screened on the basis of bronchiectasis or chronicpancreatitis, three mutations, Q1352H, E217G, and IVS8-T5 appearing with a M470V allele were associated with disease[32] . Login to comment

[hide] Misfolding of the cystic fibrosis transmembrane co... Biochemistry. 2008 Feb 12;47(6):1465-73. Epub 2008 Jan 15. Cheung JC, Deber CM
Misfolding of the cystic fibrosis transmembrane conductance regulator and disease.
Biochemistry. 2008 Feb 12;47(6):1465-73. Epub 2008 Jan 15., 2008-02-12 [PMID:18193900]

Abstract [show]
Understanding the structural basis for defects in protein function that underlie protein-based genetic diseases is the fundamental requirement for development of therapies. This situation is epitomized by the cystic fibrosis transmembrane conductance regulator (CFTR)-the gene product known to be defective in CF patients-that appears particularly susceptible to misfolding when its biogenesis is hampered by mutations at critical loci. While the primary CF-related defect in CFTR has been localized to deletion of nucleotide binding fold (NBD1) residue Phe508, an increasing number of mutations (now ca. 1,500) are being associated with CF disease of varying severity. Hundreds of these mutations occur in the CFTR transmembrane domain, the site of the protein's chloride channel. This report summarizes our current knowledge on how mutation-dependent misfolding of the CFTR protein is recognized on the cellular level; how specific types of mutations can contribute to the misfolding process; and describes experimental approaches to detecting and elucidating the structural consequences of CF-phenotypic mutations.

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145 When we examined the potential structural impact of CF-phenotypic mutations in extracellular loop 2 (ECL2) (including E217G and Q220R) in a library of wild type and mutant TM3-ECL2- TM4 hairpin constructs, we found that SDS-PAGE gel migration rates differed over a range of nearly 40% +/- the wild type position, and that decreased migration rates FIGURE 6: Disruption of helix-helix interactions by increased R-helical structure in the extracellular loop. Login to comment

[hide] Cystic fibrosis transmembrane conductance regulato... Int J Urol. 2008 Mar;15(3):270-1. Sakamoto H, Yajima T, Suzuki K, Ogawa Y
Cystic fibrosis transmembrane conductance regulator (CFTR) gene mutation associated with a congenital bilateral absence of vas deferens.
Int J Urol. 2008 Mar;15(3):270-1., [PMID:18304229]

Abstract [show]
Cystic fibrosis transmembrane conductance regulator (CFTR) gene mutations associated with cystic fibrosis have been reported to be rare in Japanese patients with congenital bilateral absence of vas deferens (CBAVD). A 28-year-old Japanese male was referred for infertility. Vas deferens and epididymis were not palpable bilaterally. Semen analyses showed azoospermia with volumes below 2.0 ml. Serum follicle-stimulating hormone value was slightly elevated. Seminal fructose concentration was also very low. Scrotal ultrasonography showed absence of the bodies and tails of the right and left epididymides. Imaging studies showed cystic dysplasia of the right seminal vesicle and agenesis of the left seminal vesicle. A CFTR gene mutation of I556V was found. Recent studies show that prevalence of CFTR gene mutation in Japanese CBAVD patients may be approximately equal to that of the Caucasian population. Genetic counselling may be recommended for any couple attempting assisted reproduction technology when the man has CBAVD.

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28 Anzai et al. showed, using polymerase chain reaction (PCR) amplification single-strand confirmation polymorphism analysis and direct sequencing to analyze all 27 exons of the CFTR gene in 19 Japanese CBAVD patients, that three rare CFTR gene mutations (W216X, G1349S, Q1352H) were found in seven CFTR alleles of five patients, and IVS8-5T was positive in 11 alleles of 11 patients.2 Namely, 58% of the 19 CBAVD patients had at least one mutated CFTR allele.2 Moreover, three (5.7%) of 53 normal individuals had a missense mutation in one of their CFTR genes (E217G in 1, and Q1352H in 2).2 CFTR gene mutations may be frequently associated with Japanese CBAVD patients. Login to comment

[hide] Atypical cystic fibrosis and CFTR-related diseases... Clin Rev Allergy Immunol. 2008 Dec;35(3):116-23. Paranjape SM, Zeitlin PL
Atypical cystic fibrosis and CFTR-related diseases.
Clin Rev Allergy Immunol. 2008 Dec;35(3):116-23., [PMID:18493878]

Abstract [show]
Cystic fibrosis (CF), which is among the most common life-shortening recessive illnesses, is caused by mutations of the CF transmembrane conductance regulator (CFTR) and typically involves chronic infection and progressive obstruction of the respiratory tract as well as pancreatic exocrine insufficiency. Disease severity, to some extent, correlates with organ sensitivity to CFTR dysfunction and to the amount of functional protein, which is influenced by the type of mutation. Atypical CF represents approximately 2% of affected individuals, and includes cases presenting in adolescence or adulthood with pancreatic exocrine sufficiency, normal or borderline sweat chloride concentrations, or with a single predominant clinical feature. This review briefly describes diagnostic methods and phenotypic characteristics of classic and atypical CF, as well as CFTR-related diseases, conditions in which mutated CFTR may contribute to the pathogenesis but do not strictly fit established diagnostic criteria.

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64 Determination of the transepithelial nasal potential difference has been beneficial in establishing a CF Table 1 Mutations, sites, and molecular consequences associated with either an atypical presentation of CF respiratory disease or pancreatic sufficiency or late-onset pancreatic insufficiency (http:// www.genet.sickkids.on.ca) Mutation Site Consequence Atypical presentation M1210I Exon 19 Met to Ile at 1210 S1455X Exon 24 Ser to Stop at 1455 1811+18G→A Intron 11 mRNA splicing defect L346P Exon 7 Leu to Pro at 346 Y161D Exon 4 Tyr to Asp at 161 R31C Exon 2 Arg to Cys at 31 I752S Exon 13 Ile to Ser at 752 2811G/T Exon 15 Sequence variation Pancreatic sufficiency or late-onset pancreatic insufficiency R600G Exon 13 Arg to Gly at 600 D1152H Exon 18 Asp to His at 1152 Y89C Exon 3 Tyr to Cys at 89 R117H Exon 4 Arg to His at 117 D110E Exon 4 Asp to Glu at 110 296 + 3insT Intron 2 mRNA splicing defect E217G Exon 6a Glu to Gly at 217 V392G Exon 8 Val to Gly at 392 N1088D Exon 17b Asn to Asp at 1088 S737F Exon 13 Missense 1716+1G→A Intron 10 mRNA splicing defect R334W Exon 7 Arg to Trp at 334 R347P Exon 7 Arg to Pro at 347 A455E Exon 9 Ala to Glu at 455 P574H Exon 12 Pro to His at 574 3850-3T→G Intron 19 mRNA splicing defect diagnosis in many atypical cases. Login to comment

[hide] The L441P mutation of cystic fibrosis transmembran... J Korean Med Sci. 2010 Jan;25(1):166-71. Epub 2009 Dec 26. Gee HY, Kim CK, Kim SW, Lee JH, Kim JH, Kim KH, Lee MG
The L441P mutation of cystic fibrosis transmembrane conductance regulator and its molecular pathogenic mechanisms in a Korean patient with cystic fibrosis.
J Korean Med Sci. 2010 Jan;25(1):166-71. Epub 2009 Dec 26., [PMID:20052366]

Abstract [show]
Cystic fibrosis (CF) is an autosomal recessive disorder usually found in populations of white Caucasian descent. CF is caused by mutations in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene. A 5-yr-old Korean girl was admitted complaining of coughing and greenish sputum. Chest radiographs and computed tomographic (CT) scan revealed diffuse bronchiectasis in both lungs. The patient had chronic diarrhea and poor weight gain, and the abdominal pancreaticobiliary CT scan revealed atrophy of the pancreas. Finally, CF was confirmed by the repeated analysis of the quantitative pilocarpine iontophoresis test. The chloride concentration of sweat samples taken from both forearms of the pateint was an average of 88.7 mM/L (normal value <40 mM/L). After a comprehensive search for mutations in the CFTR gene, the patient was found to carry the non-synonymous L441P mutation in one allele. Molecular physiologic analysis of the L441P mutation of CFTR revealed that the L441P mutation completely abolished the CFTR Cl(-) channel activity by disrupting proper protein folding and membrane trafficking of CFTR protein. These results confirmed the pathogenicity of the L441P mutation of CFTR circulating in the Korean population. The possibility of CF should be suspected in patients with chronic bronchiectasis, although the frequency of CF is relatively rare in East Asia.

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141 The heterozygote frequency of mild mutations, such as E217G and Q1352H, was estimated to 0.51% in the Korean population. Login to comment

[hide] Identification of the second CFTR mutation in pati... Asian J Androl. 2010 Nov;12(6):819-26. Epub 2010 Jul 26. Giuliani R, Antonucci I, Torrente I, Grammatico P, Palka G, Stuppia L
Identification of the second CFTR mutation in patients with congenital bilateral absence of vas deferens undergoing ART protocols.
Asian J Androl. 2010 Nov;12(6):819-26. Epub 2010 Jul 26., [PMID:20657600]

Abstract [show]
Congenital bilateral absence of vas deferens (CBAVD) is a manifestation of the mildest form of cystic fibrosis (CF) and is characterized by obstructive azoospermia in otherwise healthy patients. Owing to the availability of assisted reproductive technology, CBAVD patients can father children. These fathers are at risk of transmitting a mutated allele of the CF transmembrane conductance regulator (CFTR) gene, responsible for CF, to their offspring. The identification of mutations in both CFTR alleles in CBAVD patients is a crucial requirement for calculating the risk of producing a child with full-blown CF if the female partner is a healthy CF carrier. However, in the majority of CBAVD patients, conventional mutation screening is not able to detect mutations in both CFTR alleles, and this difficulty hampers the execution of correct genetic counselling. To obtain information about the most represented CFTR mutations in CBAVD patients, we analysed 23 CBAVD patients, 15 of whom had a single CFTR mutation after screening for 36 mutations and the 5T allele. The search for the second CFTR mutation in these cases was performed by using a triplex approach: (i) first, a reverse dot-blot analysis was performed to detect mutations with regional impact; (ii) next, multiple ligation-dependent probe amplification assays were conducted to search for large rearrangements; and (iii) finally, denaturing high-performance liquid chromatography was used to search for point mutations in the entire coding region. Using these approaches, the second CFTR mutation was detected in six patients, which increased the final detection rate to 60.8%.

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58 INNO-LiPA CFTR19 INNO-LiPA CFTR17 INNO-LiPA CFTR Italian regional [delta]F508 621+1G>T 1259insA G542X 3849+10kbC>T 4016insT N1303K 2183AA>G 4382delA W1282X 394delTT 852del22 G551D 2789+5G> A R1162X D579G 1717-1G>A 3659delC G1244E R553X R117H G1349D CFTRdele2,3 (21 kb) R334W I502T [delta]I507 R347P L1065P 711+1G>T G85E R1158X 3272-26A>G 3905insT 1078delT T338I R560T A455E S549R(A>C) 1898+1G>A S1251N 2143delA 711+5G>A 991del5 I148T E60X D1152H 3199del6 3120+1G>A 2184delA 1898+3A>G, R1070Q Q552X Poli-T tract variations R1066H R347H 621+3A>G R334Q E217G Abbreviation: CFTR, cystic fibrosis transmembrane conductance regulator. Login to comment

[hide] Association between cystic fibrosis transmembrane ... Yonsei Med J. 2010 Nov;51(6):912-7. Kim KW, Lee JH, Lee MG, Kim KH, Sohn MH, Kim KE
Association between cystic fibrosis transmembrane conductance regulator gene mutations and susceptibility for childhood asthma in Korea.
Yonsei Med J. 2010 Nov;51(6):912-7., [PMID:20879059]

Abstract [show]
PURPOSE: Classic cystic fibrosis is now known part of cystic fibrosis transmembrane conductance regulator (CFTR)-related disorders. These include a wide spectrum, from multi-system disorders, such as cystic fibrosis, to mono-symptomatic conditions, such as chronic pancreatitis or congenital bilateral absence of the vas deferens. However, respiratory disease is considered typical for the multi system disorder, cystic fibrosis, and is the major cause of morbidity and mortality. The purpose of this study was to evaluate the potential effects of CFTR gene mutations in Korean children with asthma. MATERIALS AND METHODS: We selected 14 mutations identified in Korea and each of the 48 children with and without asthma were genotyped for the case-control study. RESULTS: No significant differences were found in genotype and allele frequencies of the 9 polymorphisms observed between the non-asthma and asthma groups. In a haplotype determination based on a Bayesian algorithm, 8 haplotypes were assembled in the 98 individuals tested. However, we also did not find any significant differences in haplotype frequencies between the non-asthma and asthma groups. CONCLUSION: We have concluded that this study did not show any evidence in support of providing that CFTR genetic variations significantly contribute to the susceptibility of asthma in Korean children.

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53 CFTR Genetic Variations Analyzed in This Study Name Nucleotide change Exon Consequence Reference - 8G / C G to C at 125 5` UTR sequence variation 9 Q98R A to G at 425 Exon 4 Gln to Arg at 98 8 I125T T to C at 506 Exon 4 Ile to Thr at 125 9 E217G A to G at 782 Exon 6a Glu to Gly at 217 9 Q220X C to T at 790 Exon 6a Gln to Stop at 220 7, 8 A309A C or G at 1059 Exon 7 Sequence variation 9 TG repeat TG10-13 IVS 8 Splicing 9 T repeat T5-9 IVS 8 Splicing 9 M470V A or G at 1540 Exon 10 Met to Val at 470 9 I556V A to G at 1798 Exon 11 Ile to Val at 556 9 T854T T to G at 2694 Exon 14a Sequence variation 9 Q1291X C to T at 4003 Exon 20 Gln to Stop at 1291 9 Q1352H G to C at 4188 Exon 22 Gln to His at 1352 9 R1453W C to T at 4489 Exon 24 Arg to Trp at 1453 9 CFTR,cysticfibrosistransmembraneconductanceregulator. Login to comment
69 Frequency of CFTR Genetic Variations in Non-Asthma and Asthma Group Variants Non-asthma (n) Asthma (n) p value* - 8G / C G / G 39 37 0.466 G / C 8 11 C / C 1 0 E217G A / A 48 46 0.247 A / G 0 2 M470V A / A 8 10 0.858 A / G 25 23 G / G 15 15 I556V A / A 42 45 0.276 A / G 4 3 T854T T / T 15 16 0.639 T / G 26 22 G / G 7 10 Q1352H G / G 46 46 0.383 G / C 2 2 R1453W C / C 47 46 0.500 C / T 0 1 Microsatellite TG repeat (IVS 8)� W / W� 10 12 0.119 W / M 27 18 M / M 10 18 T repeat (IVS 8) 5 / 7 2 1 0.141 6 / 7 0 1 7 / 7 44 42 7 / 9 1 4 CFTR,cysticfibrosistransmembraneconductanceregulator. Login to comment
75 However the background haplotype for ∆F508,27 which accounts for 66% of worldwide cystic fibrosis, is very rare in the Korean population.11 Besides, genetic variants at Q1352H or E217G were found to be associated with bronchiectasis and/or chronic pancreatitis in the Korean population.11 In particular, nonsynonymous Q1352H and E217G mutations in the M470 background caused a 60-80% reduction in CFTR-dependent Cl- currents and HCO3-transport activities. Login to comment

[hide] Hepatobiliary transport. J Hepatol. 2000;32(1 Suppl):3-18. Kullak-Ublick GA, Beuers U, Paumgartner G
Hepatobiliary transport.
J Hepatol. 2000;32(1 Suppl):3-18., [PMID:10728790]

Abstract [show]
The alterations of hepatobiliary transport that occur in cholestasis can be divided into primary defects, such as mutations of transporter genes or acquired dysfunctions of transport systems that cause defective canalicular or cholangiocellular secretion, and secondary defects, which result from biliary obstruction. The dysfunction of distinct biliary transport systems as a primary cause of cholestasis is exemplified by the genetic defects in progressive familial intrahepatic cholestasis or by the direct inhibition of transporter gene expression by cytokines. In both, the hepatocellular accumulation of toxic cholephilic compounds causes multiple alterations of hepatocellular transporter expression. In addition, lack of specific components of bile caused by a defective transporter, as in the case of mdr2/MDR3 deficiency, unmasks the toxic potential of other components. The production of bile is critically dependent upon the coordinated regulation and function of sinusoidal and canalicular transporters, for instance of Na+-taurocholate cotransporting polypeptide (NTCP) and bile salt export pump (BSEP). Whereas the downregulation of the unidirectional sinusoidal uptake system NTCP protects the hepatocyte from further intracellular accumulation of bile salts, the relative preservation of canalicular BSEP expression serves to uphold bile salt secretion, even in complete biliary obstruction. Conversely, the strong downregulation of canalicular MRP2 (MRP, multidrug resistance protein) in cholestasis forces the hepatocyte to upregulate basolateral efflux systems such as MRP3 and MRP1, indicating an inverse regulation of basolateral and apical transporters The regulation of hepatocellular transporters in cholestasis adheres to the law of parsimony, since many of the cellular mechanisms are pivotally governed by the effect of bile salts. The discovery that bile salts are the natural ligand of the farnesoid X receptor has shown us how the major bile component is able to regulate its own enterohepatic circulation by affecting transcription of the genes critically involved in transport and metabolism.

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223 How- 10 ever, the major pathogenetic factor in estradiol-induced cholestasis appears to be inhibition of Bsep transport function by the cholestatic estrogen metabolite estradiol- 17fl-D-glucuronide (E217G) (152). Login to comment
224 E217G, which is an mrp2 substrate (153), probably trans-inhibits Bsep function from within the canalicular lumen, since mrp2-deficient rat strains that are unable to secrete E217G into the bile canaliculus do not develop cholestasis (154). Login to comment
225 In accordance with these observations, it has been shown that Bsep function is inhibited neither in canalicular plasma membrane vesicles of mrp2-deficient GY/TR rat liver, nor in Bsep expressing membrane vesicles from baculovirus-infected Sf9 cells (68), indicating that intact mrp2 function is a prerequisite for the development of E217G induced cholestasis. Login to comment

[hide] Comparative mutant analysis of Arabidopsis ABCC-ty... Plant Cell Physiol. 2008 Apr;49(4):557-69. Epub 2008 Mar 6. Frelet-Barrand A, Kolukisaoglu HU, Plaza S, Ruffer M, Azevedo L, Hortensteiner S, Marinova K, Weder B, Schulz B, Klein M
Comparative mutant analysis of Arabidopsis ABCC-type ABC transporters: AtMRP2 contributes to detoxification, vacuolar organic anion transport and chlorophyll degradation.
Plant Cell Physiol. 2008 Apr;49(4):557-69. Epub 2008 Mar 6., [PMID:18325934]

Abstract [show]
The enormous metabolic plasticity of plants allows detoxification of many harmful compounds that are generated during biosynthetic processes or are present as biotic or abiotic toxins in their environment. Derivatives of toxic compounds such as glutathione conjugates are moved into the central vacuole via ATP-binding cassette (ABC)-type transporters of the multidrug resistance-associated protein (MRP) subfamily. The Arabidopsis genome contains 15 AtMRP isogenes, four of which (AtMRP1, 2, 11 and 12) cluster together in one of two major phylogenetic clades. We isolated T-DNA knockout alleles in all four highly homologous AtMRP genes of this clade and subjected them to physiological analysis to assess the function of each AtMRP of this group. None of the single atmrp mutants displayed visible phenotypes under control conditions. In spite of the fact that AtMRP1 and AtMRP2 had been described as efficient ATP-dependent organic anion transporters in heterologous expression experiments, the contribution of three of the AtMRP genes (1, 11 and 12) to detoxification is marginal. Only knockouts in AtMRP2 exhibited a reduced sensitivity towards 1-chloro-2,4-dinitrobenzene, but not towards other herbicides. AtMRP2 but not AtMRP1, 11 and 12 is involved in chlorophyll degradation since ethylene-treated rosettes of atmrp2 showed reduced senescence, and AtMRP2 expression is induced during senescence. This suggests that AtMRP2 is involved in vacuolar transport of chlorophyll catabolites. Vacuolar uptake studies demonstrated that transport of typical MRP substrates was reduced in atmrp2. We conclude that within clade I, only AtMRP2 contributes significantly to overall organic anion pump activity in vivo.

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13 Abbreviations: ABC, ATP-binding cassette transporter; ABRC, Arabidopsis Biological Resource Center; Bn-NCC-1, Brassica napus non-fluorescent chlorophyll catabolite 1; BPT1, bile pigment transporter 1; BSA, bovine serum albumin; CDNB, 1-chloro-2,4-dinitrobenzene; CFTR, cystic fibrosis transmembrane conductance regulator; DNB, dinitrobenzene; DTT, dithiothreitol; E217G, 17b-estradiol 17-(b-D-glucuronide); GFP, green fluorescent protein; GS, glutathione; GS-X, glutathione conjugate; GUS, b-glucuronidase; LTC4, leukotriene C4; NASC, Nottingham Arabidopsis Stock Center; MRP, (ABCC) multidrug resistance-associated protein; NBD, nucleotide-binding domain; ORF, open reading frame; PAR, photosynthetically active radiation; PGP, P-glycoprotein; RT-PCR, reverse transcription-PCR; TMD, transmembrane domain; TWD1, twisted dwarf 1; UTR, untranslated region; YCF1, Yeast cadmium factor 1. Login to comment
49 Interestingly, AtMRP1 and AtMRP2 but not members of clade II physically interact with the C-terminal tetratricopeptide repeat domain of the membrane-bound FKBP-like immunophilin TWD1 (TWISTED DWARF1) which agrees with a modulation of vacuolar metolachlor-GS and 17b-estradiol 17-(b-D-glucuronide) (E217G) uptake by recombinant TWD1 (Geisler et al. 2004). Login to comment
112 Col-6 mesophyll vacuoles possessed ATP-dependent transport activities for the organic glucuronide E217G which appears in the mammalian liver during steroid degradation, as already demonstrated for a variety of mono-and dicotyledonous plants (Table 1; Klein et al. 2006). Login to comment
113 Efficient inhibition of vacuolar E217G uptake by vanadate confirmed that transport occurred via an ABC transporter. Login to comment
114 In contrast, E217G uptake was significantly reduced to about 70% when vacuoles were isolated from atmrp2-1 plants (P50.05). Login to comment
122 Previous studies using isolated rye and barley vacuoles and the characterization of transport activities of heterologously expressed AtMRP2 demonstrated that transport of glucuronides such as E217G is strongly stimulated by DNB-GS (Klein et al. 1998, Liu et al. 2001). Login to comment
153 Thus, either a clade II AtMRP has the Table 1 ATP-dependent uptake of [3 H]estradiol 17-b-D-glucuronide (E217G) or [3 H]leukotriene C4 (LTC4) into rosette leaf mesophyll vacuoles isolated from atmrp2-1 and its corresponding wild type (Col-6/gla1) in the presence of MgATP Line/treatment Transport into isolated mesophyll vacuoles [pmol Â (ml vacuolar volume Â min)À1 ] Substrate E217G LTC4 Col-6/gla1 0.82 Æ 0.10 (100%) 308 Æ 71 (100%) Col-6/gla1 þ 1 mM vanadate 0.15 Æ 0.03 (18%) 18 Æ 6 (6%) Col-6/gla1 þ 0.2 mM DNB-GS 0.65 Æ 0.07 (79%) n.d. atmrp2-1 0.55 Æ 0.10 (67%)Ã 175 Æ 24 (57%)Ã atmrp2-1 þ 1 mM vanadate 0.10 Æ 0.05 (12%)Ã 11 Æ 5 (3.5%)Ã atmrp2-1 þ 0.2 mM DNB-GS 0.43 Æ 0.08 (53%)Ã n.d. In the absence of AtMRP2, ATP-dependent organic anion transport rates are reduced to 60-70%. Login to comment
186 Reduced vacuolar transport activities in the atmrp2-1 mutant The ATP-dependent, vanadate-sensitive vacuolar transport activities for the glutathione conjugate LTC4 and the glucuronide E217G-two model substrates for MRP/ABCC-type transporters-were reduced to 60 and 70%, respectively, when mesophyll vacuoles isolated from atmrp2-1 were compared with the corresponding wild type vacuoles (Table 1). Login to comment
193 Interestingly, Arabidopsis mesophyll vacuoles did not display indications of cross-stimulation of organic anion transport since transport of E217G was not stimulated by the GS-conjugate DNB-GS in our experiments. Login to comment
261 Unless indicated otherwise, for each time point and condition five polyethylene tubes (0.4 ml capacity) were prepared as follows: 70 ml of medium G [22% (v/v) Percoll, 0.4 M sorbitol, 30 mM KCl, 20 mM HEPES-KOH, pH 7.2, 0.12% (w/v) BSA, 1 mM DTT] containing 1 mM MgSO4 (without ATP) or 6 mM MgSO4 and 5 mM ATP (with ATP), and substrates (0.2 mCi of [3 H]E217G and 10 mM E217G or 20 nCi (1.27 nM) of [3 H]LTC4) and further substances as indicated were placed on the bottom of the tube. Login to comment

[hide] Regulation of expression of the multidrug resistan... J Pharmacol Exp Ther. 2002 Aug;302(2):407-15. Gerk PM, Vore M
Regulation of expression of the multidrug resistance-associated protein 2 (MRP2) and its role in drug disposition.
J Pharmacol Exp Ther. 2002 Aug;302(2):407-15., [PMID:12130697]

Abstract [show]
The multidrug resistance protein 2 (MRP2; ABCC2) is an ATP-binding cassette transporter accepting a diverse range of substrates, including glutathione, glucuronide, and sulfate conjugates of many endo- and xenobiotics. MRP2 generally performs excretory or protective roles, and it is expressed on the apical domain of hepatocytes, enterocytes of the proximal small intestine, and proximal renal tubular cells, as well as in the brain and the placenta. MRP2 is regulated at several levels, including membrane retrieval and reinsertion, translation, and transcription. In addition to transport of conjugates, MRP2 transports cancer chemotherapeutics, uricosurics, antibiotics, leukotrienes, glutathione, toxins, and heavy metals. Several mutagenesis studies have described critical residues for substrate binding and various naturally occurring mutations that eliminate MRP2 expression or function. MRP2 is important clinically as it modulates the pharmacokinetics of many drugs, and its expression and activity are also altered by certain drugs and disease states.

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51 Site-directed mutagenesis of Lys to Met (K325M) and Arg to Leu (R586L) of rat Mrp2 markedly reduced transport of DNP-SG and leukotriene C4, without affecting transport capacity of model glucuronide and sulfate conjugates yet increased the affinity for transport of E217G (Ito et al., 2001c). Login to comment
55 In other recent studies, a highly conserved tryptophan residue, Trp1246 , in the last transmembrane segment (TM17) of MSD3 of MRP1 has been shown to be essential for transport of E217G (Ito et al., 2001b). Login to comment
56 Mutation of the analogous Trp1254 of MRP2 showed this amino acid to be essential for MRP2 transport of methotrexate; nonconservative substitutions (Ala, Cys) eliminated E217G transport, whereas conservative substitutions (Tyr, Phe) were without effect (Ito et al., 2001a). Login to comment
87 Finally, E217G causes a rapid inhibition of bile flow and retrieval of Mrp2 into intracellular sites; bile flow recovers spontaneously and is followed by the exocytic insertion of Mrp2 into the canalicular membrane (Mottino et al., 2002). Login to comment
129 The transport efficiency (Vmax/Km) of substrates for rat and human MRP2 have been ranked as follows: leukotriene C4 Ͼ leukotriene D4 Ͼ 2,4DNP-SG Ͼ monoglucuronosyl bilirubin Ͼ E217G Ͼ taurolithocholate sulfate Ͼ oxidized glutathione (Keppler et al., 1997). Login to comment
157 When the cells were grown to confluent monolayers, transcellular transport of bromosulfophthalein, leukotriene C4, E217G, dehydroepiandrosterone sulfate, Fluo-3, and rifampin was higher in double-transfected (OATP8/MRP2) cells than in single-transfected (MRP2 only) cells (Cui et al., 2001). Login to comment
158 Similarly, Sasaki et al. (2002) coexpressed OATP2 (SLC21A6) and MRP2 in MDCKII cells and demonstrated vectorial transport of E217G, leukotriene C4, and taurolithocholate sulfate but not dehydroepiandrosterone sulfate or estrone-3-sulfate. Login to comment
164 Similarly, E217G inhibited Bsep in Bsep/ Mrp2 coexpressing Sf9 membrane vesicles but not in Bsep- alone expressing Sf9 membrane vesicles (Stieger et al., 2000). Login to comment
172 Similarly, although E217G is a substrate for MDR1 expressed in Sf9 insect cells, there is no measurable E217G-stimulated ATPase activity (Huang et al., 1998). Login to comment

[hide] The Arabidopsis thaliana ABC transporter AtMRP5 co... EMBO J. 2001 Apr 17;20(8):1875-87. Gaedeke N, Klein M, Kolukisaoglu U, Forestier C, Muller A, Ansorge M, Becker D, Mamnun Y, Kuchler K, Schulz B, Mueller-Roeber B, Martinoia E
The Arabidopsis thaliana ABC transporter AtMRP5 controls root development and stomata movement.
EMBO J. 2001 Apr 17;20(8):1875-87., [PMID:11296221]

Abstract [show]
In the present study, we investigated a new member of the ABC transporter superfamily of Arabidopsis thaliana, AtMRP5. AtMRP5 encodes a 167 kDa protein and exhibits low glutathione conjugate and glucuronide conjugate transport activity. Promotor- beta-glucuronidase fusion constructs showed that AtMRP5 is expressed mainly in the vascular bundle and in the epidermis, especially guard cells. Using reverse genetics, we identified a plant with a T-DNA insertion in AtMRP5 (mrp5-1). mrp5-1 exhibited decreased root growth and increased lateral root formation. Auxin levels in the roots of mrp5-1 plants were increased. This observation may indicate that AtMRP5 works as an auxin conjugate transporter or that mutant plants are affected in ion uptake, which may lead to changes in auxin concentrations. Experiments on epidermal strips showed that in contrast to wild type, the sulfonylurea glibenclamide had no effect on stomatal opening in mrp5-1 plants. This result strongly suggests that AtMRP5 may also function as an ion channel regulator.

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60 ATP-dependent uptake of oestradiol-17-(b-D-glucuronide) (E217G) and a rye ¯avonoid glucuronide was reported for vacuoles from rye and barley (Klein et al., 1998, 2000). Login to comment
63 The yeast mutant transformed with AtMRP5 was able to transport E217G when compared with the empty vector control. Login to comment
65 Reduced glutathione, oxidized glutathione and dinitrobenzene glutathione (DNB-GS) had no effect on E217G uptake. Login to comment
66 However, the AtMRP5-dependent transport activity of E217G was severely in¯uenced by other organic anions such as oestradiol-3-sulfate, the natural ¯avone-glu-curonide luteolin-7-O-diglucuronide-4'-O-glucuronide, glycocholate and the sulfonylurea glibenclamide. Login to comment
109 Due to the variability of the uptake activities in different preparations, uptake rates were standardized to 100%, which corresponds to 7±25 pmol E217G/mg protein/min. Login to comment
206 In addition to GS-Xs, AtMRP5 was also able to transport the model glucuronide E217G, a substance produced during the catabolism of steroids in animals but not known in plants. Login to comment
229 The vacuolar transport rate for E217G and ¯avone glucuronides was strongly increased in the presence of the conjugate DNB-GS. Login to comment

[hide] Differential sensitivity of plant and yeast MRP (A... FEBS Lett. 2003 Nov 6;554(1-2):23-9. Forestier C, Frangne N, Eggmann T, Klein M
Differential sensitivity of plant and yeast MRP (ABCC)-mediated organic anion transport processes towards sulfonylureas.
FEBS Lett. 2003 Nov 6;554(1-2):23-9., [PMID:14596908]

Abstract [show]
The role of ATP-binding cassette (ABC) proteins such as multidrug resistance-associated proteins (MRPs) is critical in drug resistance in cancer cells and in plant detoxification processes. Due to broad substrate spectra, specific modulators of these proteins are still lacking. Sulfonylureas such as glibenclamide are used to treat non-insulin-dependent diabetes since they bind to the sulfonylurea receptor. Glibenclamide also inhibits the cystic fibrosis transmembrane conductance regulator, p-glycoprotein in animals and guard cell ion channels in plants. To investigate whether this compound is a more general blocker of ABC transporters the sensitivity of ABC-type transport processes across the vacuolar membrane of plants and yeast towards glibenclamide was evaluated. Glibenclamide inhibits the ATP-dependent uptake of beta-estradiol 17-(beta-D-glucuronide), lucifer yellow CH, and (2',7'-bis-(2-carboxyethyl)-5-(and-6-)carboxyfluorescein. Transport of glutathione conjugates into plant but not into yeast vacuoles was drastically reduced by glibenclamide. Thus, irrespective of the homologies between plant, yeast and animal MRP transporters, specific features of plant vacuolar MRPs with regard to sensitivity towards sulfonylureas exist. Glibenclamide could be a useful tool to trap anionic fluorescent indicator dyes in the cytosol.

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31 Abbreviations: ABC, ATP-binding cassette; BCECF, (2P,7P-bis-(2-carboxyethyl)-5-(and-6-)carboxy£uorescein; BmCl, monochlorobi- mane; CFTR, cystic 'brosis transmembrane conductance regulator; DNB-GS, 2,4-dinitrobenzene glutathione; E217G, L-estradiol 17-(L-D- glucuronide); GSH, reduced glutathione; GSSG, oxidized glutathione; GS-X, glutathione conjugate; KCO, potassium channel opener; LY-CH, lucifer yellow CH; MRP, multidrug resistance-associated protein; SUR, sulfonylurea receptor FEBS 27703 23-10-03 Cyaan Magenta Geel Zwart FEBS 27703 FEBS Letters 554 (2003) 23^29 trol other membrane proteins and more speci'cally ion channels [17]. Login to comment
69 Protoplast preparations were incapable of transporting glutathione conjugates or E217G. Login to comment
92 Results It has been demonstrated that isolated vacuoles or tonoplast vesicles from various plant sources take up GS-X as well as E217G using directly energized transport systems [6,10,33]. Login to comment
107 Furthermore, cromakalim does not antagonize glucuronide transport inhibition by gli- Fig. 1. Dose-dependent inhibition of vacuolar E217G uptake into barley vacuoles by glibenclamide in the absence (circles) or presence (squares) of 0.2 mM DNB-GS. Login to comment
108 Barley mesophyll vacuoles were incubated with 11.4 nM E217G in the presence of 4 mM MgCl2, 3 mM ATP and glibenclamide at the concentrations indicated. Login to comment
113 Table 1 E¡ects of sulfonylureas and KCOs on the ATP-dependent uptake of [3 H]E217G into barley vacuoles Condition Vacuolar transport of [3 H]E217G without DNB-GS +0.2 mM DNB-GS % of MgATP-stimulated value Control 100 832 þ 320 +150 WM glibenclamide (glib) 12.9 þ 12.7 36 þ 29 +0.1 mM tolbutamide 88.1 þ 7.4 638 þ 9 +1 mM tolbutamide 81.6 þ 5.2 599 þ 86 +10 WM cromakalim (crom) 90.2 þ 2.2 500 þ 15 +100 WM crom 63 þ 9.5 518 þ 13 +1 WM SR47063 87.9 þ 18.53 536 þ 17 +10 WM SR47063 35.6 þ 20.7 298 þ 61 +10 WM RP49356 79.2 þ 4.67 595 þ 11 +100 WM RP49356 63.1 þ 2.62 373 þ 29 +150 WM glib+100 WM crom 2.8 þ 3.9 49.4 þ 5 +15 WM glib+100 WM crom 64.1 þ 5.2 429 þ 56 Barley mesophyll vacuoles were incubated with 11.4 nM E217G in the presence of 4 mM MgCl2, 3 mM ATP, in the absence or presence of DNB-GS as a stimulating agent and diverse pharmacological compounds interfering with CFTR and SUR. Login to comment
119 Vacuoles isolated from an Arabidopsis cell culture exhibited a directly energized transport systems for DNB-GS and E217G. Login to comment
121 Thus, the glibenclamide sensitivity of directly energized transport systems for DNB-GS and E217G is not species-speci'c in plants. Login to comment
123 Glibenclamide selectively inhibits the vacuolar deposition of anionic £uorescent dyes in barley vacuoles Since glibenclamide was a very e/cient inhibitor of E217G but not DNB-GS transport, we investigated its action on the vacuolar accumulation of di¡erent negatively charged £uorescent probes used as substrates for ABC-type transport processes. Login to comment
126 Vacuolar LY-CH uptake was stimulated by 0.2 mM DNB-GS up to about 920% of the control suggesting a similar interaction with GS-Xs as shown for E217G. Login to comment
127 Again, glibenclamide strongly inhibited the DNB-GS-stimulated LY-CH uptake into barley vacuoles (from about 920 to about 5%; Fig. 2) suggesting a comparable pharmacology as shown for E217G. Login to comment
132 In contrast to LY-CH and E217G, the vacuolar uptake of BCECF was not stimulated by di¡erent GS-X and was inhibited by oxidized glutathione (GSSG) (Table Table 3 BCECF uptake into barley vacuoles is mediated by an ABC-type transporter Treatment Uptake of BCECF % of MgATP-stimulated value 3ATP 0.1 þ 0.1 +MgATP (control) 100 +MgATP +1 mM vanadate 5.1 þ 7.3 +MgATP +0.1 WM ba'lomycin A1 103.9 þ 26.0 +MgATP +5 mM NH4Cl 129.4 þ 24.7 +MgATP +150 WM glibenclamide 0.0 þ 2.2 +MgATP +3 mM GSSG 55.1 þ 13.4 +MgATP +3 mM GSH 89.8 þ 15.4 +MgATP +0.2 mM DNB-GS 115.5 þ 6.2 +MgATP +0.2 mM Decyl-GS 93.8 þ 19.8 Isolated barley vacuoles were incubated with 1 mM MgSO4 (3ATP) or 3 mM ATP and 4 mM MgSO4 (+MgATP), 25 WM BCECF and further compounds as indicated. Login to comment
196 This hypothesis is in accordance with the fact that the CFTR chloride channel is blocked by glibenclamide and taurocholate or E217G [42]. Login to comment
207 Swanson et al. [34] suggested the presence of a vacuolar ABC-type transport system for BCECF in bar- Fig. 5. Dose-dependent glibenclamide inhibition of the ATP-stimulated uptake of E217G (closed circles) and DNB-GS (open circles) in yeast vacuoles. Login to comment
208 Yeast vacuoles isolated from the W303-A1 strain were incubated with 10 WM E217G or 15 WM DNB-GS in the presence of 5 mM MgATP. Login to comment
197 This hypothesis is in accordance with the fact that the CFTR chloride channel is blocked by glibenclamide and taurocholate or E217G [42]. Login to comment
209 Yeast vacuoles isolated from the W303-A1 strain were incubated with 10 WM E217G or 15 WM DNB-GS in the presence of 5 mM MgATP. Login to comment

[hide] Casein kinase 2alpha regulates multidrug resistanc... Mol Pharmacol. 2012 Sep;82(3):488-99. doi: 10.1124/mol.112.078295. Epub 2012 Jun 13. Stolarczyk EI, Reiling CJ, Pickin KA, Coppage R, Knecht MR, Paumi CM
Casein kinase 2alpha regulates multidrug resistance-associated protein 1 function via phosphorylation of Thr249.
Mol Pharmacol. 2012 Sep;82(3):488-99. doi: 10.1124/mol.112.078295. Epub 2012 Jun 13., [PMID:22695718]

Abstract [show]
We have shown previously that the function of Ycf1p, yeast ortholog of multidrug resistance-associated protein 1 (MRP1), is regulated by yeast casein kinase 2alpha (Cka1p) via phosphorylation at Ser251. In this study, we explored whether casein kinase 2alpha (CK2alpha), the human homolog of Cka1p, regulates MRP1 by phosphorylation at the semiconserved site Thr249. Knockdown of CK2alpha in MCF7-derived cells expressing MRP1 [MRP1 CK2alpha(-)] resulted in increased doxorubicin sensitivity. MRP1-dependent transport of leukotriene C(4) and estradiol-17beta-d-glucuronide into vesicles derived from MRP1 CK2alpha(-) cells was decreased compared with MRP1 vesicles. Moreover, mutation of Thr249 to alanine (MRP1-T249A) also resulted in decreased MRP1-dependent transport, whereas a phosphomimicking mutation (MRP1-T249E) led to dramatic increase in MRP1-dependent transport. Studies in tissue culture confirmed these findings, showing increased intracellular doxorubicin accumulation in MRP1 CK2alpha(-) and MRP1-T249A cells compared with MRP1 cells. Inhibition of CK2 kinase by 2-dimethylamino-4,5,6,7-tetrabromo-1H-benzimidazole resulted in increased doxorubicin accumulation in MRP1 cells, but not in MRP1 CK2alpha(-), MRP1-T249A, or MRP1-T249E cells, suggesting that CK2alpha regulates MRP1 function via phosphorylation of Thr249. Indeed, CK2alpha and MRP1 interact physically, and recombinant CK2 phosphorylates MRP1-derived peptide in vitro in a Thr249-dependent manner, whereas knockdown of CK2alpha results in decreased phosphorylation at MRP1-Thr249. The role of CK2 in regulating MRP1 was confirmed in other cancer cell lines where CK2 inhibition decreased MRP1-mediated efflux of doxorubicin and increased doxorubicin cytotoxicity. This study supports a model in which CK2alpha potentiates MRP1 function via direct phosphorylation of Thr249.

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178 Transport of E217G also shows a trend to be reduced; however, it did not reach statistical significance (Fig. 3F). Login to comment

[hide] Mutations in the cystic fibrosis transmembrane con... J Cyst Fibros. 2012 Jul;11(4):316-23. doi: 10.1016/j.jcf.2012.01.005. Epub 2012 Apr 6. Li H, Wen Q, Li H, Zhao L, Zhang X, Wang J, Cheng L, Yang J, Chen S, Ma X, Wang B
Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) in Chinese patients with congenital bilateral absence of vas deferens.
J Cyst Fibros. 2012 Jul;11(4):316-23. doi: 10.1016/j.jcf.2012.01.005. Epub 2012 Apr 6., [PMID:22483971]

Abstract [show]
BACKGROUND: Genetic testing of the cystic fibrosis transmembrane conductance (CFTR) gene is currently performed in patients with congenital bilateral absence of vas deferens (CBAVD). This study was conducted to investigate the role of mutations in the CFTR gene in CBAVD-dependent male infertility. METHODS: 73 Chinese patients diagnosed with CBAVD were studied. The entire coding regions and splice sites of 27 exons of the CFTR gene were sequenced in 146 chromosomes from the 73 CBAVD patients. Screening was carried out using PCR, gel electrophoresis and DNA sequencing to identify novel variants of the entire coding regions and boundaries of the 27 exons. RESULTS: Five novel nonsynonymous mutations, three novel splice site mutations and one deletion were identified by sequencing. Apart from the novel variants, we also found 19 previously reported mutations and polymorphism sites. Thirty-four patients (46.57%) had the 5T variant (6 homozygous and 28 heterozygous) and in two of them it was not associated with any detectable mutation of the CFTR gene. All potential pathogenic mutations are not contained in the 1000 Genome Project database. In total, the present study identified 30 potential pathogenic variations in the CFTR gene, 9 of which had not previously been described. CONCLUSIONS: Most patients with CBAVD have mutations in the CFTR gene. 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 CBAVD patients and will facilitate the development of more sensitive CFTR mutation screening.

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77 Lastly, we have observed previously reported mutations and polymorphisms (p.E217G, p.R347H, p.V470M, p.R553X, p.I556V, p.T854T, p.G970D, p.P1290P, p.Q1352H, p.Q1643Q, 744-5delGATT, IVS8-T5) (Supplementary Table 1). Login to comment
119 △F508 R117H Mutation genotypes IVS8-Tn n (%) Two mutations detected Neg Neg I556V/I556V 7T/7T 1(1.3) Neg Neg I556V/1209+2 G-C 5T/7T 1(1.3) Neg Neg I556V/726delATT 5T/5T 1(1.3) Neg Neg I556V/- 5T/5T 1(1.3) Neg Neg I556V/- 5T/7T 1(1.3) Neg Neg G970D/- 5T/7T 1(1.3) Neg Neg C592F/- 5T/5T 1(1.3) Neg Neg 1209+1 G-C/- 5T/7T 1(1.3) Neg Neg R553X/- 5T/7T 1(1.3) Neg Neg Q1352H/- 5T/7T 1(1.3) Neg Neg S485C/- 5T/7T 1(1.3) Neg Neg A357T/- 5T/7T 1(1.3) Neg Neg E217G/- 5T/7T 1(1.3) Neg Neg R347H/- 5T/7T 1(1.3) Neg Neg G451K/- 5T/7T 1(1.3) Neg Neg L558S/- 5T/7T 1(1.3) Neg Neg 3635delT/Q1352H 7T/7T 1(1.3) Neg Neg A1136T/G970D 7T/7T 1(1.3) Neg Neg 870-1 G-C/- 5T/7T 1(1.3) Neg Neg 520-2 A-G/- 5T/7T 1(1.3) Neg Neg R419I/- 5T/7T 1(1.3) Neg Neg C491F/Q1643Q 7T/7T 1(1.3) Neg Neg Q1352H/- 5T/7T 1(1.3) Neg Neg R851X/- 5T/7T 1(1.3) Neg Neg P750L/G970D 7T/7T 1(1.3) One mutation detected Neg Neg -/- 5T/7T 2(2.7) Neg Neg -/- 5T/7T 3(4.1) Neg Neg -/- 5T/7T 5(6.8) Neg Neg -/- 5T/5T 2(2.7) Neg Neg -/- 5T/5T 1(1.3) Neg Neg G970D/- 7T/7T 2(2.7) Neg Neg D993Y/- 7T/7T 1(1.3) Neg Neg I556V/- 7T/7T 1(1.3) Neg Neg T388R/- 7T/7T 1(1.3) No mutation detected Neg Neg -/- 7T/7T 8(10.9) Neg Neg -/- 7T/7T 15(20.5) Neg Neg -/- 7T/9T 2(2.7) Neg Neg -/- 7T/7T 4(5.5) Neg: Negative. Login to comment
76 Lastly, we have observed previously reported mutations and polymorphisms (p.E217G, p.R347H, p.V470M, p.R553X, p.I556V, p.T854T, p.G970D, p.P1290P, p.Q1352H, p.Q1643Q, 744-5delGATT, IVS8-T5) (Supplementary Table 1). Login to comment
118 b3;F508 R117H Mutation genotypes IVS8-Tn n (%) Two mutations detected Neg Neg I556V/I556V 7T/7T 1(1.3) Neg Neg I556V/1209+2 G-C 5T/7T 1(1.3) Neg Neg I556V/726delATT 5T/5T 1(1.3) Neg Neg I556V/- 5T/5T 1(1.3) Neg Neg I556V/- 5T/7T 1(1.3) Neg Neg G970D/- 5T/7T 1(1.3) Neg Neg C592F/- 5T/5T 1(1.3) Neg Neg 1209+1 G-C/- 5T/7T 1(1.3) Neg Neg R553X/- 5T/7T 1(1.3) Neg Neg Q1352H/- 5T/7T 1(1.3) Neg Neg S485C/- 5T/7T 1(1.3) Neg Neg A357T/- 5T/7T 1(1.3) Neg Neg E217G/- 5T/7T 1(1.3) Neg Neg R347H/- 5T/7T 1(1.3) Neg Neg G451K/- 5T/7T 1(1.3) Neg Neg L558S/- 5T/7T 1(1.3) Neg Neg 3635delT/Q1352H 7T/7T 1(1.3) Neg Neg A1136T/G970D 7T/7T 1(1.3) Neg Neg 870-1 G-C/- 5T/7T 1(1.3) Neg Neg 520-2 A-G/- 5T/7T 1(1.3) Neg Neg R419I/- 5T/7T 1(1.3) Neg Neg C491F/Q1643Q 7T/7T 1(1.3) Neg Neg Q1352H/- 5T/7T 1(1.3) Neg Neg R851X/- 5T/7T 1(1.3) Neg Neg P750L/G970D 7T/7T 1(1.3) One mutation detected Neg Neg -/- 5T/7T 2(2.7) Neg Neg -/- 5T/7T 3(4.1) Neg Neg -/- 5T/7T 5(6.8) Neg Neg -/- 5T/5T 2(2.7) Neg Neg -/- 5T/5T 1(1.3) Neg Neg G970D/- 7T/7T 2(2.7) Neg Neg D993Y/- 7T/7T 1(1.3) Neg Neg I556V/- 7T/7T 1(1.3) Neg Neg T388R/- 7T/7T 1(1.3) No mutation detected Neg Neg -/- 7T/7T 8(10.9) Neg Neg -/- 7T/7T 15(20.5) Neg Neg -/- 7T/9T 2(2.7) Neg Neg -/- 7T/7T 4(5.5) Neg: Negative. Login to comment

[hide] CFTR gene mutations in Japanese individuals with c... J Cyst Fibros. 2003 Mar;2(1):14-8. Anzai C, Morokawa N, Okada H, Kamidono S, Eto Y, Yoshimura K
CFTR gene mutations in Japanese individuals with congenital bilateral absence of the vas deferens.
J Cyst Fibros. 2003 Mar;2(1):14-8., [PMID:15463840]

Abstract [show]
Congenital bilateral absence of the vas deferens (CBAVD) is a monosymptomatic disease confined to the male reproductive system with similarity to the phenotype of cystic fibrosis (CF), and mutations in the CFTR gene are highly prevalent in Caucasian CBAVD patients. While CF is very rare in Japan, CBAVD is not. Our previous study demonstrated high prevalence of the 5T allele in the CFTR gene in Japanese CBAVD patients. We analyzed whole exons of the CFTR gene in 19 CBAVD patients and 53 normal individuals using polymerase chain reaction amplification-single strand conformation polymorphism analysis and direct sequencing. Three missense mutations (W216X, G1349S, Q1352H) were found in seven CFTR alleles, and the 5T allele was positive in 11 of 38 CFTR patient alleles. Consequently, 47% of CFTR chromosomes in the patients were affected, and 11 individuals (58%) had at least one mutated CFTR allele. In contrast, three of 53 normal individuals (5.7%) had a missense mutation in one of the CFTR genes, but no 5T allele was detected (both P<0.0001). Mutations of the CFTR gene are closely associated with Japanese patients with CBAVD.

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54 In contrast, in 53 normal individuals similarly tested, E217G was found in one chromosome and Q1352H was in two, but no other mutations were detected (data not shown). Login to comment

[hide] CFTR mutations spectrum and the efficiency of mole... PLoS One. 2014 Feb 26;9(2):e89094. doi: 10.1371/journal.pone.0089094. eCollection 2014. Zietkiewicz E, Rutkiewicz E, Pogorzelski A, Klimek B, Voelkel K, Witt M
CFTR mutations spectrum and the efficiency of molecular diagnostics in Polish cystic fibrosis patients.
PLoS One. 2014 Feb 26;9(2):e89094. doi: 10.1371/journal.pone.0089094. eCollection 2014., [PMID:24586523]

Abstract [show]
Cystic fibrosis (CF) is caused by mutations in the cystic fibrosis transmembrane regulator gene (CFTR). In light of the strong allelic heterogeneity and regional specificity of the mutation spectrum, the strategy of molecular diagnostics and counseling in CF requires genetic tests to reflect the frequency profile characteristic for a given population. The goal of the study was to provide an updated comprehensive estimation of the distribution of CFTR mutations in Polish CF patients and to assess the effectiveness of INNOLiPA_CFTR tests in Polish population. The analyzed cohort consisted of 738 patients with the clinically confirmed CF diagnosis, prescreened for molecular defects using INNOLiPA_CFTR panels from Innogenetics. A combined efficiency of INNOLiPA CFTR_19 and CFTR_17_TnUpdate tests was 75.5%; both mutations were detected in 68.2%, and one mutation in 14.8% of the affected individuals. The group composed of all the patients with only one or with no mutation detected (109 and 126 individuals, respectively) was analyzed further using a mutation screening approach, i.e. SSCP/HD (single strand conformational polymorphism/heteroduplex) analysis of PCR products followed by sequencing of the coding sequence. As a result, 53 more mutations were found in 97 patients. The overall efficiency of the CF allele detection was 82.5% (7.0% increase compared to INNOLiPA tests alone). The distribution of the most frequent mutations in Poland was assessed. Most of the mutations repetitively found in Polish patients had been previously described in other European populations. The most frequent mutated allele, F508del, represented 54.5% of Polish CF chromosomes. Another eight mutations had frequencies over 1%, 24 had frequencies between 1 and 0.1%; c.2052-2053insA and c.3468+2_3468+3insT were the most frequent non-INNOLiPA mutations. Mutation distribution described herein is also relevant to the Polish diaspora. Our study also demonstrates that the reported efficiency of mutation detection strongly depends on the diagnostic experience of referring health centers.

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No. Sentence Comment
51 Three of them (R352Q, Q359R and D1152H) were in a compound heterozygosity with F508del, six (E217G, I506, V562L, G723V, D924N and L967S) had no accompanying mutation in trans. Login to comment
71 Exon / intron (legacy) Exon / intron (Ensembl) Protein change SVM value cDNA (HGVS nomenclature) gDNA (cDNA +132 bp) Number of PL CF chromosomes Reference a Mutations in trans Pathogenic mutations 1 1 L15Ffs10X c.43delC 175delC 1 CFMDB 1717-1G.A 2 2 G27V 21.92 c.80G.T 212G.T 1 Novel F508del 2 2 S18RfsX16 c.54-5940_273 +10250del21kb exon2,3del21kb 66 IL19 various CF mutations i2 i2 IVS2_Donor c.164+1G.A 296+1G.A 3 CFMDB various CF mutations 3 3 G85E 22.61 c.254G.A 386G.A 1 IL17 unknown 3 3 E60X c.178G.T 310G.T 0 IL17 x 3 3 L88IfsX22 c.262_263delTT 394delTT 0 IL17 x 4 4 E92K 21.92 c.274G.A 406G.A 2 CFMDB c.164+1G.A; c.2051- 2AA.G 4 4 L101X c.302T.G 434T.G 1 CFMDB c.3717+12191C.T 4 4 K114IfsX5 c.341_353del13bp 473del13bp 1 Novel F508del 4 4 R117H 20.35 c.350G.A 482G.A 5 IL17 F508del; 2x unknown 4 4 R117C 22.07 c.349C.T 481C.T 2 CFMDB S1206X;1x unknown 4 4 L137_L138insT c.412_413insACT L138ins 1 CFMDB F508del 4 4 R153I 22.61 c.458G.T 590G.T 2 Novel F508del; c.3527delC i4 i4 IVS4_Donor c.489+1G.T 621+1G.T 5 IL17 F508del; c.489+1G.T 5 5 L165X c.494T.A 626T.A 1 Novel F508del i5 i5 IVS5_Donor c.579+1G.T 711+1G.T 0 IL19 x i5 i5 IVS5_Donor c.579+3A.G 711+3A.G 2 CFMDB 2,3del21kb; c.2052-3insA i5 i5 IVS5_Donor c.579+5G.A 711+5G.A 0 IL17 x 7 8 F311L 20.90 c.933C.G 965C.G 2 CFMDB 2x F508 7 8 G314R 20.58 c.940G.A 1072G.A 4 CFMDB various CF mutations 7 8 F316LfsX12 c.948delT 1078delT 1 IL17 unkown 7 8 R334W 22.41 c.1000C.T 1132C.T 6 IL17 various CF mutations 7 8 I336K 22.07 c.1007T.A 1139T.A 2 CFMDB 2,3de21kb; F508del 7 8 R347P 22.27 c.1040G.C 1172G.C 11 IL17 various CF mutations i7 i8 IVS8_Donor c.1116+2T.A 1248+2T.A 1 Novel Q1412X 9 10 A455E 22.61 c.1364C.A 1496C.A 0 IL17 x i9 i10 IVS10_Donor c.1392+1G.A 1524+1G.A 1 CFMDB c.3816-7delGT 10 11 S466X c.1397C.G 1529C.G 1 CFMDB G542X 10 11 I507del c.1519_1521delATC 1651delATC 2 IL19 F508del 10 11 F508del c.1521_1523delCTT 1654delCTT 805 IL19 various CF mutations i10 i11 IVS11_Acceptor c.1585-1G.A 1717-1G.A 27 IL19 various CF mutations 11 12 G542X c.1624G.T 1756G.T 25 IL19 various CF mutations 11 12 G551D 21.24 c.1624G.T 1756G.T 5 IL19 various CF mutations 11 12 Q552X c.1654C.T 1786C.T 0 IL19 x 11 12 R553X c.1657C.T 1789C.T 14 IL19 various CF mutations 11 12 R560T 21.92 c.1679G.C 1811G.C 0 IL19 x i12 i13 IVS13_Donor c.1766+1G.A 1898+1G.A 6 IL19 various CF mutations i12 i13 IVS13_Donor c.1766+1G.C 1898+1G.C 1 CFMDB F508del 13 14 H620P 21.73 c.1859A.C 1991A.C 1 CFMDB F508del 13 14 R668C//G576A 21.61//1.73 c.2002C.T//c.1727G.C 2134C.T// 1859G.C 5 b CFMDB// rs1800098 c.1585-1G.A; 4 unknown 13 14 L671X c.2012delT 2143delT 27 IL17 various CF mutations 13 14 K684SfsX38 c.2051_2052delAAinsG 2183AA.G 10 IL17 various CF mutations 13 14 K684NfsX38 c.2052delA 2184delA 0 IL17 x 13 14 Q685TfsX4 c.2052_2053insA 2184insA 15 CFMDB various CF mutationsc , 1 unknown Table 2. Cont. Exon / intron (legacy) Exon / intron (Ensembl) Protein change SVM value cDNA (HGVS nomenclature) gDNA (cDNA +132 bp) Number of PL CF chromosomes Reference a Mutations in trans 13 14 L732X c.2195T.G 2327T.G 1 CFMDB F508del 14A 15 R851X c.2551C.T 2683C.T 3 CFMDB various CF mutations 14A 15 I864SfsX28 c.2589_2599del11bp 2721del11bp 2 CFMDB F508del; 2,3del21kb i14B i16 IVS16_Donor c.2657+2_2657+3insA 2789+2insA 1 CFMDB F508del i14B i16 IVS16_Donor c.2657+5G.A 2789+5G.A 0 IL17 unkown 15 17 Y919C 21.02 c.2756A.G 2888A.G 1 CFMDB unknown 15 17 H939HfsX27 c.2817_2820delTACTC 2949delTACTC 1 Novel unkown i15 i17 IVS17_Donor c.2908+3A.C 3040+3A.C 1 Novel F508del i16 i18 IVS18_Donor c.2988+1G.A 3120+1G.A 0 IL19 x 17A 19 I1023_V1024del c.3067_3072delATAGTG 3199del6 0 IL19 x i17A i19 IVS19 c.3140-26A.G 3272-26A.G 9 IL19 various CF mutations 17B 20 L1065R 21.90 c.3194T.G 3326T.G 1 CFMDB F508del 17B 20 Y1092X c.3276C.A 3408C.A 1 CFMDB R334W i18 i21 IVS21_Donor c.3468+2_3468+3insT 3600+2insT 11 CFMDB various CF mutationsd , 1 unknown 18 21 E1126EfsX7 c.3376_3379delGAAG 3508delGAAG 1 Novel F508del 19 22 R1158X c.3472C.T 3604C.T 2 CFMDB F508del; R553X 19 22 R1162X c.3484C.T 3616C.T 1 IL17 F508del 19 22 L1177SfsX15 c.3528delC 3659delC 4 IL17 various CF mutations 19 22 S1206X c.3617C.A 3749C.A 1 CFMDB R117C i19 i22 IVS22 c.3717+12191C.T 3849+10kbC.T 58 IL17 various CF mutations 20 23 G1244R 22.62 c.3730G.C 3862G.C 1 CFMDB F508del 20 23 S1251N 22.28 c.3752G.A 3884G.A 0 IL19 x 20 23 L1258FfsX7 c.3773_3774insT 3905insT 0 IL19 x 20 23 V1272VfsX28 c.3816_3817delGT 3944delGT 1 CFMDB c.1392+1G.A 20 23 W1282X c.3846G.A 3978G.A 9 IL19 various CF mutations 21 24 N1303K 22.62 c.3909C.G 4041C.G 18 IL19 various CF mutations 22 25 V1327X c.3979delG 4111delG 1 Novel F508del 22 25 S1347PfsX13 c.4035_4038dupCCTA c.4167dupCCTA 1 CFMDB 2,3del21kb 23 26 Q1382X c.4144C.T 4276C.T 1 CFMDB F508del 23 26 Q1412X c.4234C.T 4366C.T 2 CFMDB F508del; c.1116+2T.A i23 i26 IVS26_Donor c.4242+1G.T 4374+1G.T 1 CFMDB F508del Sequence changes of uncertain pathogenic effect, tentatively counted as mutations 6A 6 E217G 0.30 c.650A.G 782A.G 1 CFMDB; rs1219109046 unknown 7 8 R352Q 20.01 c.1055G.A 1187G.A 1 CFMDB; rs121908753 F508del 7 8 Q359R 0.33 c.1076A.G 1208A.G 1 CFMDB F508del i8 i9 IVS9 c.1210-12T5_1210- 34_35 (TG)12 1332-12Tn_- 34TGm 6 CFMDB F508del; 3x unknown i8 i9 IVS9 c.1210-12T5_1210- 34_35 (TG)13 1332-12Tn_- 34TGm 2 CFMDB 2143delT; 1x unknown i8 i9 IVS9 c.1210-12T8 1332-12Tn 1 Novel unknown 10 11 I506V 20.21 c.1516A.G 1648A.G 1 CFMDB; rs1800091 unknown 12 13 V562L 0.79 c.1684G.C 1816G.C 1 CFMDB; rs1800097 unknown 13 14 G723V 0.44 c.2168G.T 2300G.T 1 CFMDB; rs200531709 unknown 15 17 D924N 0.03 c.2770G.A 2902G.A 1 CFMDB; rs201759207 unknown patient with F508del on another allele) was not supported by the SVM value (+0.35); the patient was PS and had ambiguous chloride values (45, 64 and 83 mmol/L). 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[hide] Functional characteristics of L1156F-CFTR associat... Am J Physiol Gastrointest Liver Physiol. 2015 Aug 15;309(4):G260-9. doi: 10.1152/ajpgi.00015.2014. Epub 2015 Jun 18. Kondo S, Fujiki K, Ko SB, Yamamoto A, Nakakuki M, Ito Y, Shcheynikov N, Kitagawa M, Naruse S, Ishiguro H
Functional characteristics of L1156F-CFTR associated with alcoholic chronic pancreatitis in Japanese.
Am J Physiol Gastrointest Liver Physiol. 2015 Aug 15;309(4):G260-9. doi: 10.1152/ajpgi.00015.2014. Epub 2015 Jun 18., [PMID:26089335]

Abstract [show]
Although cystic fibrosis is rare in Japanese, measurement of sweat Cl(-) has suggested mild dysfunction of cystic fibrosis transmembrane conductance regulator (CFTR) in some patients with chronic pancreatitis. In the present study, we have investigated the association of CFTR variants and chronic pancreatitis in Japanese and the functional characteristics of a Japanese- and pancreatitis-specific CFTR variant, L1156F. Seventy patients with alcoholic chronic pancreatitis, 18 patients with idiopathic chronic pancreatitis, and 180 normal subjects participated. All exons and their boundaries and promoter region of the CFTR gene were sequenced. Human embryonic kidney-293 cells were transfected with three CFTR variants (M470V, L1156F, and M470V+L1156F), and the protein expression was examined. Xenopus laevis oocytes were injected with the CFTR variants, and bicarbonate (HCO3 (-)) transport activity was examined. CFPAC-1 cells were transfected with the CFTR variants and Cl(-)/HCO3 (-) exchange activity was examined. Six variants (E217G, I556V, M470V, L1156F, Q1352H, and R1453W) were identified in the coding region of the CFTR gene. Cystic fibrosis-causing mutations were not found. The allele frequencies of L1156F and Q1352H in alcoholic chronic pancreatitis (5.0 and 7.9%) were significantly (P < 0.01) higher than those in normal subjects (0.6 and 1.9%). L1156F was linked with a worldwide CFTR variant, M470V. Combination of M470V and L1156F significantly reduced CFTR expression to approximately 60%, impaired CFTR-mediated HCO3 (-)/Cl(-) transport activity to 50-60%, and impaired CFTR-coupled Cl(-)/HCO3 (-) exchange activity to 20-30%. The data suggest that the Japanese-specific CFTR variant L1156F causes mild dysfunction of CFTR and increases the risk of alcoholic chronic pancreatitis in Japanese.

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Sentences [show]
No. Sentence Comment
11 Six variants (E217G, I556V, M470V, L1156F, Q1352H, and R1453W) were identified in the coding region of the CFTR gene. Login to comment
57 Six CFTR variants, c.650Ab0e;G, p.Glu217Gly (E217G); c.1666Ab0e;G, p.Ile556Val (I556V); M470V; L1156F; Q1352H; and R1453W, were detected. Login to comment
132 Six variants (E217G, I556V, M470V, L1156F, Q1352H, and R1453W) were identified in coding regions of the CFTR gene (Table 2). Login to comment
135 The allele frequencies of E217G, I556V, and M470V were not different among groups. Login to comment
141 The allele frequencies of polymorphisms in the coding regions of CFTR gene ACP ICP NS n 140 36 360 E217G (exon 6a) Glu 137 (97.9) 36 (100) 354 (98.3) Gly 3 (2.1) 0 (0) 6 (1.7) M470V (exon 10) Met 60 (42.9) 14 (38.9) 143 (39.7) Val 80 (57.1) 22 (61.1) 217 (60.3) I556V (exon 11) Ile 138 (98.6) 36 (100) 348 (96.7) Val 2 (1.4) 0 (0) 12 (3.3) L1156F (exon 18) Leu 133 (95.0) 35 (97.2) 358 (99.4) Phe 7 (5.0)* 1 (2.8) 2 (0.6) Q1352H (exon 22) Gln 129 (92.1) 35 (97.2) 353 (98.1) His 11 (7.9)* 1 (2.8) 7 (1.9) R1453W (exon 24) Arg 138 (98.6) 32 (88.9) 353 (98.1) Trp 2 (1.4) 4 (11.1)* 7 (1.9) Values are no. Login to comment