ABCMdb

ABCC7 p.Gly85Val

ClinVar:	c.254G>A , p.Gly85Glu D , Pathogenic c.254G>T , p.Gly85Val ? , not provided
CF databases:	c.254G>A , p.Gly85Glu D , CF-causing ; CFTR1: This mutation was detected in family #26, a French Canadian family classified as PI. This Gly to Glu change is associated with a group IIb haplotype. The mutation destroys a Hinfl site. The PCR product derived from the 3i-5 and 3i-3 primers is cleaved by this enzyme into 3 fragments, 172, 105, and 32 bp, respectively, for the normal sequence; a fragment of 277 bp would be present for the mutant sequence. They analyzed 54 CF chromosomes, 8 from group II and 50 normal chromosomes, 44 from group II and did not find another eample of G85E. c.254G>T , p.Gly85Val (CFTR1) ? , The patient with G85V carried G542X on the other chromosome, and presented mild phenotype with PI.
Predicted by SNAP2:	A: D (95%), C: D (95%), D: D (95%), E: D (59%), F: D (95%), H: D (95%), I: D (95%), K: D (95%), L: D (95%), M: D (95%), N: D (95%), P: D (95%), Q: D (95%), R: D (95%), S: D (95%), T: D (95%), V: D (95%), W: D (95%), Y: D (95%),
Predicted by PROVEAN:	A: N, C: D, D: D, E: D, F: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: N, T: D, V: D, W: D, Y: D,

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[hide] N-terminal CFTR missense variants severely affect ... Hum Mutat. 2008 May;29(5):738-49. Gene GG, Llobet A, Larriba S, de Semir D, Martinez I, Escalada A, Solsona C, Casals T, Aran JM
N-terminal CFTR missense variants severely affect the behavior of the CFTR chloride channel.
Hum Mutat. 2008 May;29(5):738-49., [PMID:18306312]

Abstract [show]
Over 1,500 cystic fibrosis transmembrane conductance regulator (CFTR) gene sequence variations have been identified in patients with cystic fibrosis (CF) and related disorders involving an impaired function of the CFTR chloride channel. However, detailed structure-function analyses have only been established for a few of them. This study aimed evaluating the impact of eight N-terminus CFTR natural missense changes on channel behavior. By site-directed mutagenesis, we generated four CFTR variants in the N-terminal cytoplasmic tail (p.P5L, p.S50P, p.E60K, and p.R75Q) and four in the first transmembrane segment of membrane-spanning domain 1 (p.G85E/V, p.Y89C, and p.E92K). Immunoblot analysis revealed that p.S50P, p.E60K, p.G85E/V, and p.E92K produced only core-glycosylated proteins. Immunofluorescence and whole cell patch-clamp confirmed intracellular retention, thus reflecting a defect of CFTR folding and/or trafficking. In contrast, both p.R75Q and p.Y89C had a glycosylation pattern and a subcellular distribution comparable to the wild-type CFTR, while the percentage of mature p.P5L was considerably reduced, suggesting a major biogenesis flaw on this channel. Nevertheless, whole-cell chloride currents were recorded for all three variants. Single-channel patch-clamp analyses revealed that the channel activity of p.R75Q appeared similar to that of the wild-type CFTR, while both p.P5L and p.Y89C channels displayed abnormal gating. Overall, our results predict a major impact of the CFTR missense variants analyzed, except p.R75Q, on the CF phenotype and highlight the importance of the CFTR N-terminus on channel physiology.

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156 Confocal images from representative xy sections taken from1of 3 independent experiments show the subcellular distribution of wild-type CFTR (WT), p.F508del mutant (F508del), and variants p.S50P (S50P), p.E60K (E60K), p.G85E (G85E), p.G85V (G85V), p.E92K (E92K), p.P5L (P5L), p.R75Q (R75Q), and p.Y89C (Y89C). Login to comment
180 B: Recordings fromvariants p.S50P (S50P), p.E60K (E60K), p.G85E (G85E), p.G85V (G85V), and p.E92K (E92K) (superimposed recordings). Login to comment
40 The eight CFTR variants included in this study: p.P5L, p.S50P, p.E60 K, p.R75Q, p.G85E, p.G85V, p.Y89C, and p.E92K (Fig. 1A) were generated by oligonucleotide-directed mutagenesis in pCMVCFTRNot6.2wt using the QuickChangeTM XL-Site-Directed Mutagenesis Kit (Stratagene, La Jolla, CA) according to the manufacturer`s instructions (see Supplementary Table S1 for a detailed description of the mutagenesis primers employed; available online at http://www.interscience.wiley.com/jpages/1059-7794/supp mat). Login to comment
102 RESULTS Description and Cross-Species Analysis of Natural N-Terminus CFTR Variants We chose eight naturally occurring sequence variants, four located across the N-terminal CFTR tail (p.P5L, p.S50P, p.E60K, and p.R75Q), and four within the first segment of MSD1 (p.G85E, p.G85V, p.Y89C, and p.E92 K) (Fig. 1A; Table 1). Login to comment
110 In contrast, variants p.S50P, p.E60K, p.G85E, p.G85V, and p.E92K, produced only higher mobility band B proteins suggesting that, like the p.F508del mutant, the resulting misfolded channels are retained and degraded in the cytoplasm (Fig. 2A). Login to comment
118 B: Alignment of the N-terminus (amino acids 1 to 100) of the CFTR protein derived from di¡erent species.The sequences derived from human (Homo sapiens, Gen- BankNM_000492), mouse (Mus musculus,GenBankNM_021050), Norway rat (Rattusnovergicus,GenBankNM_031506), European rabbit (Oryctolagus cuniculus, GenBank NM_001082716), cow (Bos taurus, GenBank NM_174018), sheep (Ovis aries, GenBank NM_001009781), African clawed frog (Xenopus laevis, GenBank X65256), and spiny dog'sh (Squalus acanthias, GenBank M83785) were aligned using the ClustalW multiple sequence alignment program.The amino acid residue a¡ected by each of the variants analyzed (p.P5L, p.S50P, p.E60K, p.R75Q, p.G85E, p.G85V, p.Y89C, and p.E92K) is indicated in bold. Login to comment
122 Similarly, variants p.S50P, p.E60K, p.G85V, p.G85E, and p.E92K displayed a yellow colocalization pattern clearly compatible with retention of the anomalous CFTR proteins within the intracellular compartments and no detection of PM staining (Fig. 3). Login to comment
130 Likewise, none of the five variants (p.S50P, p.E60K, p.G85E, p.G85V, and p.E92K) in which severe misprocessing was previously demonstrated (Figs. 2 and 3), was able to generate cAMP-activated currents (Fig. 4B). Login to comment
133 Genotype^Phenotype Correlation in the N-Terminal CFTR MissenseVariants Under StudyÃ Missense varianta Phenotype Second allele (number of patients)b p.P5L CF p.F508del (1), p.P205S (1) p.S50P CBAVD p.F508del (1), p.E115del (1) p.E60K CF p.G542X (1), p.I507del (1) p.R75Q HT p.F508del (3), p.E725K (1) B p.R347H (1), p.R75Q (1), n.i. (4) Br c.1584G4A (2), c.1210-7_1210-6delTT (1), n.i.(3) NT p.F508del (1) CP c.1584G4A (1), n.i. (3) MI n.i. (1) CUAVD n.i. (2) OZ n.i. (2) Normal p.R75Q (1), c.2052_2053insA (1), n.i. (1) p.G85E CF p.F508del (8), p.G542X (2), p.I507del (1), c.580-1G4T (1), p.G85E (1), c.1477_ 1478delCA (1) CBAVD p.G576A (1) HT p.L997F (1),WT (1) p.G85V CF p.F508del (2), p.G542X (2), p.Y1092X (1), c.265715G4A (1), p.A1006E, c.1210-7_1210- 6delTT (1), n.i. (1) p.Y89C CF n.i. (1)c p.E92K CF p.F508del (2), p.Q890X (1), p.L206W (1) CBAVD c.1210-7_1210-6delTT (1) ÃThe recommendations for mutation nomenclature (www.hgvs.org/mutnomen/) were used to name CFTR gene sequence variations at both the nucleotide level and the protein level. Login to comment
213 Four of the variants (p.P5L, p.S50P, p.E60 K, and p.R75Q) are localized within the cytosolic N-terminal tail, and the remaining four (p.G85E, p.G85V, p.Y89C, and p.E92K) are embedded in three positions within the first transmembrane segment (TM1) of MSD1. Login to comment
215 Accordingly, using three different approaches (immunoblotting, immunocytochemistry, and electrophysiology) we found that 5 (p.S50P, p.E60K, p.G85E, p.G85V, and p.E92K) out of the 8 variants failed to mature, showing an analogous behavior than the most common F508del mutation. Login to comment
220 Moreover, although both p.G85E and p.G85V, similarly to p.E92K, do not appear to affect TM1 topology [Xiong et al., 1997], they are also localized within or adjacent to the bilayer. Login to comment

[hide] Independent contribution of common CFTR variants t... Pancreas. 2010 Mar;39(2):209-15. de Cid R, Ramos MD, Aparisi L, Garcia C, Mora J, Estivill X, Farre A, Casals T
Independent contribution of common CFTR variants to chronic pancreatitis.
Pancreas. 2010 Mar;39(2):209-15., [PMID:19812525]

Abstract [show]
OBJECTIVE: We have assessed whether CFTR gene has a major impact on chronic pancreatitis (CP) pathogenesis than that provided by the CFTR mutations. For this aim, we have evaluated clinical parameters, CFTR mutations, and 3 potential regulatory CFTR variants (coding single-nucleotide polymorphisms): c.1540A>G, c.2694T>G, and c.4521G>A. METHODS: CFTR gene analysis was performed in a cohort of 136 CP patients and 93 controls from Spanish population using current scanning techniques (single-strand conformation polymorphism/heteroduplex, denaturing gradient gel electrophoresis, and denaturing high-performance liquid chromatography) and direct sequencing. RESULTS: A higher frequency of CFTR mutations were observed in patients (39%) than in controls (15%; P < or = 0.001), differences being mostly attributable to the prevalence of the cystic fibrosis (CF)-causing mutations (P = 0.009). The analysis of variants has shown statistically significant differences between patients and controls for c.4521G>A (Pcorrected = 0.036). Furthermore, the multi-marker analysis revealed that the 1540A;2694G;4521A (AGA) haplotype was more prevalent in CP than controls (Pcorrected = 0.042). Remarkably, this association was unrelated to CF-causing mutations (P = 0.006). CONCLUSIONS: Our results corroborate the higher susceptibility of CF carriers to CP and, furthermore, suggest that the AGA haplotype could contribute to an increased risk in the development of CP irrespective of other CF-causing mutations.

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38 Scanning Methodology Applied in CFTR Gene Analysis Amplicon Name Fragment Size, bp Control Set (n = 93) Patient Set 1 (n = 68) Patient Set 2 (n = 68) Control Sequence Exon 1 192 SSCP/HD SSCP/HD dHPLC 125G9C Exon 2 334 SSCP/HD SSCP/HD dHPLC 296+3insT Exon 3 309 DGGE DGGE dHPLC G85V Exon 4 436 SSCP/HD SSCP/HD dHPLC R117H Exon 5 466 DGGE DGGE dHPLC R170H Exon 6a 345 SSCP/HD SSCP/HD dHPLC L206W Exon 6b 331 SSCP/HD SSCP/HD SSCP/HD TTGA 6/7 Exon 7 410 SSCP/HD SSCP/HD dHPLC R334W Exon 8 328 DGGE DGGE dHPLC 1341+28C9T Exon 9 375 DGGE DGGE DGGE 7T/9T Exon 10 493 SSCP/HD SSCP/HD SSCP/HD F508del; 1540A/A Exon 11 322 DGGE DGGE dHPLC S549R Exon 12 426 DGGE DGGE dHPLC G576A Exon 13a 532 SSCP/HD SSCP/HD dHPLC R668C Exon 13b 498 SSCP/HD SSCP/HD dHPLC I807M Exon 14a 284 DGGE DGGE DGGE 2694T9G Exon 14b 211 DGGE DGGE dHPLC 2789+5G9A Exon 15 487 DGGE DGGE dHPLC D924N Exon 16 294 SSCP/HD SSCP/HD dHPLC 3041-71G9C Exon 17a 294 SSCP/HD SSCP/HD dHPLC L997F Exon 17b 463 DGGE DGGE dHPLC 3272-26A9G Exon 18 451 DGGE DGGE dHPLC N1148K Exon 19 588 SSCP/HD SSCP/HD SSCP/HD 3601-65C9A Exon 20 471 DGGE DGGE dHPLC W1282X Exon 21 477 DGGE DGGE DGGE 4029G9A Exon 22 339 SSCP/HD SSCP/HD dHPLC Q1352H Exon 23 249 DGGE DGGE dHPLC 4374+13A9G Exon 24 362 SSCP/HD SSCP/HD SSCP/HD 4521G9A Control set, general population series analyzed; patient set 1, previous patient series reported in 2004; and patient set 2, new patient series analyzed in this study. Login to comment

[hide] Do common in silico tools predict the clinical con... Clin Genet. 2010 May;77(5):464-73. Epub 2009 Jan 6. Dorfman R, Nalpathamkalam T, Taylor C, Gonska T, Keenan K, Yuan XW, Corey M, Tsui LC, Zielenski J, Durie P
Do common in silico tools predict the clinical consequences of amino-acid substitutions in the CFTR gene?
Clin Genet. 2010 May;77(5):464-73. Epub 2009 Jan 6., [PMID:20059485]

Abstract [show]
Computational methods are used to predict the molecular consequences of amino-acid substitutions on the basis of evolutionary conservation or protein structure, but their utility in clinical diagnosis or prediction of disease outcome has not been well validated. We evaluated three popular computer programs, namely, PANTHER, SIFT and PolyPhen, by comparing the predicted clinical outcomes for a group of known CFTR missense mutations against the diagnosis of cystic fibrosis (CF) and clinical manifestations in cohorts of subjects with CF-disease and CFTR-related disorders carrying these mutations. Owing to poor specificity, none of tools reliably distinguished between individual mutations that confer CF disease from mutations found in subjects with a CFTR-related disorder or no disease. Prediction scores for CFTR mutations derived from PANTHER showed a significant overall statistical correlation with the spectrum of disease severity associated with mutations in the CFTR gene. In contrast, PolyPhen- and SIFT-derived scores only showed significant differences between CF-causing and non-CF variants. Current computational methods are not recommended for establishing or excluding a CF diagnosis, notably as a newborn screening strategy or in patients with equivocal test results.

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64 Mutations in the CFTR gene grouped by clinical category Cystic fibrosis CFTR-related disease No disease T338I D614G L320V V920L L90S M470V H199R S1251N I203M G550R P111A I148T Q1291H R560K L1388Q L183I R170H I1027T S549R D443Y P499A L1414S T908N R668C S549N A455E E1401K Q151K G27E I1234L Y563N R347P C866R S1118C P1290S R75Q A559T V520F P841R M469V E1401G P67L G85E S50Y E1409K R933G G458V G178R Y1032C R248T I980K G85V V392G L973P L137H T351S R334W I444S V938G R792G R560T R555G L1339F D1305E P574H V1240G T1053I D58G G551D L1335P I918M F994C S945L L558S F1337V R810G D1152H G1247R P574S R766M D579G W1098R H949R F200I R352Q L1077P K1351E M244K L206W M1101K D1154G L375F N1303K R1066C E528D D110Y R347H R1070Q A800G P1021S S549K A1364V V392A damaging` (is supposed to affect protein function or structure) and 'probably damaging` (high confidence of affecting protein function or structure). Login to comment

[hide] Spectrum of mutations in the CFTR gene in cystic f... Ann Hum Genet. 2007 Mar;71(Pt 2):194-201. Alonso MJ, Heine-Suner D, Calvo M, Rosell J, Gimenez J, Ramos MD, Telleria JJ, Palacio A, Estivill X, Casals T
Spectrum of mutations in the CFTR gene in cystic fibrosis patients of Spanish ancestry.
Ann Hum Genet. 2007 Mar;71(Pt 2):194-201., [PMID:17331079]

Abstract [show]
We analyzed 1,954 Spanish cystic fibrosis (CF) alleles in order to define the molecular spectrum of mutations in the CFTR gene in Spanish CF patients. Commercial panels showed a limited detection power, leading to the identification of only 76% of alleles. Two scanning techniques, denaturing gradient gel electrophoresis (DGGE) and single strand conformation polymorphism/hetroduplex (SSCP/HD), were carried out to detect CFTR sequence changes. In addition, intragenic markers IVS8CA, IVS8-6(T)n and IVS17bTA were also analyzed. Twelve mutations showed frequencies above 1%, p.F508del being the most frequent mutation (51%). We found that eighteen mutations need to be studied to achieve a detection level of 80%. Fifty-one mutations (42%) were observed once. In total, 121 disease-causing mutations were identified, accounting for 96% (1,877 out of 1,954) of CF alleles. Specific geographic distributions for the most common mutations, p.F508del, p.G542X, c.1811 + 1.6kbA > G and c.1609delCA, were confirmed. Furthermore, two other relatively common mutations (p.V232D and c.2789 + 5G > A) showed uneven geographic distributions. This updated information on the spectrum of CF mutations in Spain will be useful for improving genetic testing, as well as to facilitate counselling in people of Spanish ancestry. In addition, this study contributes to defining the molecular spectrum of CF in Europe, and corroborates the high molecular mutation heterogeneity of Mediterranean populations.

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52 Mutation 0.46-0.35 9 c.1078delT #, p.R347P # 8 p.G85V, c.621 + 1G > T #, p.S549R (T > G) #, p.R553X #, c.3849 + 10kbC > T # 7 p.R347H #, c.1812-1G > A, p.R709X 0.30-0.10 6 p.H199Y, p.P205S, 5 p.R117H #, p.G551D #, p.W1089X, p.Y1092X, CFTR50kbdel 4 c.296 + 3insT, c.1717-1G > A #, c.1949del84, c.3849 + 1G > A 3 p.E92K, c.936delTA, c.1717-8G > A, c.1341G > A, p.A561E, c.2603delT, p.G1244E, [p.D1270N; p.R74W] 2 p.Q2X, p.P5L, CFTRdele2,3, p.S50P, p.E60K, c.405 + 1G > A, c.1677delTA, p.L558S, p.G673X, p.R851X, p.Y1014C, p.Q1100P, p.M1101K, p.D1152H, CFTRdele19, p.G1244V, p.Q1281X, p.Y1381X <0,1 1 c.124del23bp, p.Q30X, p.W57X, c.406-1G > A, p.Q98R, p.E115del, c.519delT, p.L159S, c.711 + 3A > T, p.W202X, c.875 + 1G > A, p.E278del, p.W361R, c.1215delG, p.L365P, p.A399D, c.1548delG, p.K536X, p.R560G, c.1782delA, p.L571S, [p.G576A; p.R668C], p.T582R, p.E585X, c.1898 + 1G > A, c.1898 + 3A > G, c.2051delTT, p.E692X, p.R851L, c.2711delT, c.2751 + 3A > G, c.2752-26A > G, p.D924N, p.S945L, c.3121-1G > A, p.V1008D, p.L1065R, [p.R1070W; p.R668C], [p.F1074L; 5T], p.H1085R, p.R1158X, c.3659delC #, c.3667del4, c.3737delA, c.3860ins31, c.3905insT #, c.4005 + 1G > A, p.T1299I, p.E1308X, p.Q1313X, c.4095 + 2T > A, rearrangements study (n = 4) Mutations identified in CF families with mixed European origin: c.182delT, p.L1254X, c.4010del4. Login to comment

[hide] Phenotype-optimized sequence ensembles substantial... Hum Mutat. 2012 Aug;33(8):1267-74. doi: 10.1002/humu.22110. Epub 2012 May 22. Masica DL, Sosnay PR, Cutting GR, Karchin R
Phenotype-optimized sequence ensembles substantially improve prediction of disease-causing mutation in cystic fibrosis.
Hum Mutat. 2012 Aug;33(8):1267-74. doi: 10.1002/humu.22110. Epub 2012 May 22., [PMID:22573477]

Abstract [show]
Cystic fibrosis transmembrane conductance regulator (CFTR) mutation is associated with a phenotypic spectrum that includes cystic fibrosis (CF). The disease liability of some common CFTR mutations is known, but rare mutations are seen in too few patients to categorize unequivocally, making genetic diagnosis difficult. Computational methods can predict the impact of mutation, but prediction specificity is often below that required for clinical utility. Here, we present a novel supervised learning approach for predicting CF from CFTR missense mutation. The algorithm begins by constructing custom multiple sequence alignments called phenotype-optimized sequence ensembles (POSEs). POSEs are constructed iteratively, by selecting sequences that optimize predictive performance on a training set of CFTR mutations of known clinical significance. Next, we predict CF disease liability from a different set of CFTR mutations (test-set mutations). This approach achieves improved prediction performance relative to popular methods recently assessed using the same test-set mutations. Of clinical significance, our method achieves 94% prediction specificity. Because databases such as HGMD and locus-specific mutation databases are growing rapidly, methods that automatically tailor their predictions for a specific phenotype may be of immediate utility. If the performance achieved here generalizes to other systems, the approach could be an excellent tool to help establish genetic diagnoses.

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153 Of the 103 test-set mutations, two CF-neutral mutations had a score higher than one standard deviation above the mean (G550R and R933G), and two CF-causing mutations had scores lower than one standard deviation below the mean (G85V and T338I); these four mutations were consistently the most difficult to accurately rank with our method. Login to comment

[hide] Validation of high-resolution DNA melting analysis... J Mol Diagn. 2008 Sep;10(5):424-34. Epub 2008 Aug 7. Audrezet MP, Dabricot A, Le Marechal C, Ferec C
Validation of high-resolution DNA melting analysis for mutation scanning of the cystic fibrosis transmembrane conductance regulator (CFTR) gene.
J Mol Diagn. 2008 Sep;10(5):424-34. Epub 2008 Aug 7., [PMID:18687795]

Abstract [show]
High-resolution melting analysis of polymerase chain reaction products for mutation scanning, which began in the early 2000s, is based on monitoring of the fluorescence released during the melting of double-stranded DNA labeled with specifically developed saturation dye, such as LC-Green. We report here the validation of this method to scan 98% of the coding sequence of the cystic fibrosis transmembrane conductance regulator (CFTR) gene. We designed 32 pairs of primers to amplify and analyze the 27 exons of the gene. Thanks to the addition of a small GC-clamp at the 5' ends of the primers, one single melting domain and one identical annealing temperature were obtained to co-amplify all of the fragments. A total of 307 DNA samples, extracted by the salt precipitation method, carrying 221 mutations and 21 polymorphisms, plus 20 control samples free from variations (confirmed by denaturing high-performance liquid chromatography analysis), was used. With the conditions described in this study, 100% of samples that carry heterozygous mutations and 60% of those with homozygous mutations were identified. The study of a cohort of 136 idiopathic chronic pancreatitis patients enabled us to prospectively evaluate this technique. Thus, high-resolution melting analysis is a robust and sensitive single-tube technique for screening mutations in a gene and promises to become the gold standard over denaturing high-performance liquid chromatography, particularly for highly mutated genes such as CFTR, and appears suitable for use in reference diagnostic laboratories.

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51 Sequences of the Primers Used for CFTR Analysis by HRM, GC Size, Amplicon Length, Number of Positive Controls Validated for Each Exon, and Positive Controls for Routine Analysis Exon Primer Sequences GC length Amplicon length (bp) Introns Number of heterozygous- positive controls Number of homozygous- positive controls Recommended control 1 LSCFE1Fmod 5Ј-CCGCCGCCGTTGAGCGGCAGGCACC-3Ј 8 200 bp 74 4 125GϾC LSCFE1Rmod 5Ј-CCGCCGCCGGCACGTGTCTTT CCGAAGCT-3Ј 8 19 M1I 2 2i5b 5Ј-CAAATCTGTATGGAGACC-3Ј 0 194 bp 39 5 R31C 2i3Љ 5Ј-CAACTAAACAATGTACATGAAC-3Ј 0 4 296ϩ1GϾT 3 LSCFe3Fmod LSCFe3Rmod 5Ј-CGCCGTTAAGGGAAATAGGACAA CTAAAATA-3Ј 5 276 bp 44 10 2 R75Q 5Ј-CCGCCGATTCACCAGATTTCGTAGTC-3Ј 6 66 G85V 4 LSCFe4FmodC 5Ј-CCGCCGCCGCCCGTGTTGAAATT CTCAGGGT-3Ј 12 361 bp 52 14 1 R117H LSCFe4RmodC 5Ј-CCGCCGCCCACATGTACGATAC AGAATATATGTGCC-3Ј 9 26 574delA 5 LSCFE5Fmod 5Ј-CCGCCGGTTGAAATTATCTAACTTTCC-3Ј 6 201 bp 13 8 624delT LSCFE5Rmod 5Ј-CCGAACTCCGCCTTTCCAGTTGT-3Ј 3 48 711ϩ1GϾT 6a LSCF6aFmod2 5Ј-CCGCCGGGGTGGAAGAT ACAATGACACCTG-3Ј 5 317 bp 25 8 C225X LSCF6aRmod2 5Ј-CCGCCGCCGCGATGCATAGAG CAGTCCTGGTT-3Ј 11 66 L206W 6b LSCFE6bFmod 5Ј-CGCGCCGCCGGATTTAC AGAGATCAGAGAG-3Ј 10 239 bp 0 2 1 R258G LSCFE6Brmod 5Ј-CCGCCGCCGAGGTGGA GTCTACCATGA-3Ј 8 66 1001ϩ11CϾT 7 LSCFE7Fmod2 5Ј-CCGCCGCCCTCTCCCTGAATTT TATTGTTATTGTTT-3Ј 13 326 bp 7 11 1078delT LSCFE7Rmod2 5Ј-CCCGCCGCCCTATAATGCAG CATTATGGT-3Ј 10 7 1248ϩ1GϾT 8 LSCFE8Fmod 5Ј-CCGGAATGCATTAATGCTAT TCTGATTC-3Ј 4 199 bp 32 7 W401X LSCFE8Rmod 5Ј-CCCGCAGTTAGGTGTTTAG AGCAAACAA-3Ј 4 18 1249-5AϾG 9 LSCFe9Fmod2 5Ј-CCGCCGCCGGGAATTATTTGAGAA AGCAAAACA-3Ј 8 279 bp 0 3 D443Y LSCFe9Rmod2 5Ј-CCGCCGCGAAAATACCTTCCAG CACTACAAACTAGAAA-3Ј 8 57 A455E 10 LSCF10FmodD 5Ј-CGCCGTTATGGGAGAACTGG AGCCTTCAGAG-3Ј 5 275 bp 0 15 1 F508del LSCF10RmodD 5Ј-CCGCAGACTAACCGATTGAAT ATGGAGCC-3Ј 4 68 E528E 11 h11i5 5Ј-TGCCTTTCAAATTCAGATTGAGC-3Ј 0 197 bp 42 13 2 G542X 11i3ter 5Ј-ACAGCAAATGCTTGCTAGACC-3Ј 0 17 G551D 12 LSCFE12Fmod 5Ј-CGCGTCATCTACACTAGATGACCAG-3Ј 4 244 bp 43 15 G576A 1898 ϩ 1GϾALSCFE12Rmod 5Ј-CCGGAGGTAAAATGCAATCTATGATG-3Ј 3 63 13 LSCF13AFmod 5Ј-CCGCCGCCGGAGACATATTG CAATAAAGTAT-3Ј 9 38 20 I601F LSCF13ARmod 5Ј-GCCTGTCCAGGAGACAGGA GCATCTC-3Ј 2 R668C LSCF13BFmod 5Ј-CCGCCGCAATCCTAACTGAG ACCTTACACCG-3Ј 2 R668C LSCF13BRmod 5Ј-CCGCCGATCAGGTTCAGGA CAGACTGC-3Ј 3 346 bp 2184insA LSCF13CFmod 5Ј-CCGCGGTGATCAGCACTGGCCC-3Ј 6 301 bp 77 L749L LSCF13CRmod 5Ј-CCGCGCGCGCGGCCAGTTTCTTG AGATAACCTTCT-3Ј 13 259 bp V754M LSCF13DFmod 5Ј-CGTGTCACTGGCCCCTCAGGC-3Ј 1 221 bp I807M LSCF13DRmof 5Ј-CCGCCGCCGCTAATCCTATGA TTTTAGTAAAT-3Ј 9 220 bp 2622ϩ1GϾA LSCf13FFmod 5Ј-CGCGGTGCAGAAAGAAGAAAT TCAATCCTAACTG-3Ј 4 R668C LSCF13FRmod 5Ј-CCGCCGTGCCATTCATTTGT AAGGGAGTCT-3Ј 6 2184insA 14a LSCF14aFmodB 5Ј-CCGACCACAATGGTGGCAT GAAACTG-3Ј 3 239 bp 35 7 1 T854T LSCF14aRmodB 5Ј-CCGCCGACTTTAAATCCAGTAAT ACTTTACAATAGAACA-3Ј 6 7 W846X 14b LSCF14bFmod 5Ј-CCGGAGGAATAGGTGAAGAT-3Ј 2 179 bp 38 4 2752-5GϾT LSCF14bRmodb 5Ј-CCGTACATACAAACATAGTGGATT-3Ј 3 59 2789ϩ5GϾT 15 LSCFE15Fmod 5Ј-CGCGCCGTGTATTGGAAA TTCAGTAAGTAACTTTGG-3Ј 7 412 bp 33 16 T908S LSCFE15Rmod 5Ј-CCGCAGCCAGCACTGCCAT TAGAAA-3Ј 4 68 S945L (table continues) phisms that we have chosen to exclude. Login to comment

[hide] High heterogeneity for cystic fibrosis in Spanish ... Hum Genet. 1997 Dec;101(3):365-70. Casals T, Ramos MD, Gimenez J, Larriba S, Nunes V, Estivill X
High heterogeneity for cystic fibrosis in Spanish families: 75 mutations account for 90% of chromosomes.
Hum Genet. 1997 Dec;101(3):365-70., [PMID:9439669]

Abstract [show]
We have analyzed 640 Spanish cystic fibrosis (CF) families for mutations in the CFTR gene by direct mutation analysis, microsatellite haplotypes, denaturing gradient gel electrophoresis, single-strand conformation analysis and direct sequencing. Seventy-five mutations account for 90.2% of CF chromosomes. Among these we have detected seven novel CFTR mutations, including four missense (G85V, T582R, R851L and F1074L), two nonsense (E692X and Q1281X) and one splice site mutation (711+3A-->T). Three variants, two in intronic regions (406-112A/T and 3850-129T/C) and one in the coding region (741C/T) were also identified. Mutations G85V, T582R, R851L, E692X and Q1281X are severe, with lung and pancreatic involvement; 711+3A-->T could be responsible for a pancreatic sufficiency/insufficiency variable phenotype; and F1074L was associated with a mild phenotype. These data demonstrate the highest molecular heterogeneity reported so far in CF, indicating that a wide mutation screening is necessary to characterize 90% of the Spanish CF alleles.

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2 Among these we have detected seven novel CFTR mutations, including four missense (G85V, T582R, R851L and F1074L), two nonsense (E692X and Q1281X) and one splice site mutation (711+3A→T). Login to comment
4 Mutations G85V, T582R, R851L, E692X and Q1281X are severe, with lung and pancreatic involvement; 711+3A→T could be responsible for a pancreatic sufficiency/insufficiency variable phenotype; and F1074L was associated with a mild phenotype. Login to comment
13 Among these are seven novel CFTR mutations, including four missense (G85V, T582R, R851L and F1074L), two nonsense (E692X and Q1281X) and one splice site mutation (711+3A→T). Login to comment
41 New mutations G85V An abnormal DGGE pattern in exon 3 due to the nucleotide change G→T at position 386 of CFTR determines the missense mutation G85V (glycine to valine). Login to comment
43 The G85V mutation was identified in a patient who carries the G542X mutation on the maternal CF allele. Login to comment
45 At 18 years of age he presents a severe CF phenotype with pancreatic insufficiency (PI) and a FEV1 of 32%, suggesting that G85V is a severe mutation. Login to comment
56 The patient carries the 1609delCA mutation on the paternal chromo- 367 Table 2 Seven new mutations and three DNA variants in the CFTR gene (IVS intervening sequence, DGGE denaturing gradient gel electrophoresis, SSCA single-strand conformation analysis) Mutations Exon/ CFTR Haplotype IVS Detection intron domain method 8CA 17bTA 17bCA G85V E.3 TM1 17 7 17 DGGE 711+3A→T I.5 - 15 7 17 DGGE T582R E.12 NB1 18 37 13 DGGE E692X E.13 R 16 46 13 SSCA R851L E.14a - 23 21 19 DGGE F1074L E.17b - 17 31 13 DGGE Q1281X E.20 NB2 16 28 13 DGGE Variants 406-112A/T I.3 - - SSCA 3850-129T/C I.19 - - DGGE 741C/T E.6a - - SSCA Fig.1 a Multiplex denaturing gradient gel electrophoretic analysis for exons 8, 5 and 18 of the CFTR gene. Login to comment
87 However, from the information obtained here, mutations G85V, T582R, E692X and Q1281X can be considered as severe, with lung and pancreatic involvement. Login to comment
92 While microsatellite haplotypes of mutations G85V and F1074L were also associated with several other mutations, the haplotypes associated with 711+3A→T, T582R and R851L are unique in the CF population studied here, providing a useful tool for mutation analysis (Morral et al. 1996). Login to comment

[hide] Quantification of major urinary metabolites of PGE... Prostaglandins Leukot Essent Fatty Acids. 2013 Aug;89(2-3):121-6. doi: 10.1016/j.plefa.2013.06.001. Epub 2013 Jun 20. Jabr S, Gartner S, Milne GL, Roca-Ferrer J, Casas J, Moreno A, Gelpi E, Picado C
Quantification of major urinary metabolites of PGE2 and PGD2 in cystic fibrosis: correlation with disease severity.
Prostaglandins Leukot Essent Fatty Acids. 2013 Aug;89(2-3):121-6. doi: 10.1016/j.plefa.2013.06.001. Epub 2013 Jun 20., [PMID:23791427]

Abstract [show]
Cystic fibrosis transmembrane conductance (CFTR) alterations are involved in the overproduction of prostaglandins (PG) in CF in vitro. We assessed the relationship between PGE-M and PGD-M urinary metabolites of PGE2 and PGD2 and CF severity. Twenty-four controls and 35 CF patients were recruited. PGE-M and PGD-M levels were measured by liquid chromatography/mass spectrometry and results were expressed as median and 25th-75th interquartile of ng/mg creatinine (Cr). PGE-M (15.63; 9.07-43.35ng/mg Cr) and PGD-M (2.16; 1.43-3.53ng/mg Cr) concentrations were higher in CF than in controls: PGE-M, (6.63; 4.35-8.60ng/mg Cr); PGD-M (1.23; 0.96-1.54ng/mg Cr). There was no correlation between metabolite levels and spirometric values. Patients with pancreatic insufficiency (n=29) had higher PGE-M levels (19.09; 9.36-52.69ng/mg Cr) than those with conserved function (n=6) (9.61; 5.78-14.34ng/mg Cr). PGE-M levels were associated with genotype severity: mild (7.14; 5.76-8.76, n=8), moderate (16.67; 13.67-28.62ng/mg Cr, n=5) and severe (22.82; 10.67-84.13ng/mg Cr). Our study confirms the key role of CFTR in the regulation of the cyclooxygenase pathway of arachidonic acid metabolism found in in vitro studies.

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113 Mutations Mutation class Severity Number Pancreatic sufficiency (n) W128X/W128X I/I Severe 1 0 I507/Q890X I/I Severe 1 0 F508del/G542X II/I Severe 2 0 F508del/2188AA4G II/I Severe 1 0 F508del/N1303K II/I Severe 3 0 F508del/1677delTA II/I Severe 1 0 F508del/2188AA4G II/I Severe 1 0 F508del/F508del II/II Severe 10 0 F508del/Q890X II/II Severe 1 0 F508del/E1308X II/II Severe 1 0 F508del/5T-12TG II/III Moderate 2 0 G542X/G85V I/III Moderate 1 0 F508del/124del23kbp II/III Moderate 1 0 G542X/M1137V I/III Moderate 1 1 I507/L206W I/IV Mild 1 0 F508del/L206W I/IV Mild 4 2 711+1G4L206W I/IV Mild 1 1 N1303K/3272-26A4G I/IV Mild 1 1 F508del/F587I II/V Mild 1 1 n&#bc;Number. Login to comment