ABCMdb

ABCC7 p.Lys696Arg

ClinVar:	c.2087A>G , p.Lys696Arg ? , not provided
Predicted by SNAP2:	A: N (57%), C: D (59%), D: D (59%), E: N (57%), F: D (66%), G: D (71%), H: N (53%), I: D (59%), L: D (59%), M: D (53%), N: N (72%), P: D (71%), Q: D (59%), R: N (87%), S: D (63%), T: N (66%), V: D (59%), W: D (71%), Y: D (63%),
Predicted by PROVEAN:	A: N, C: D, D: N, E: N, F: D, G: N, H: N, I: D, L: D, M: N, N: N, P: N, Q: N, R: N, S: N, T: N, V: D, W: D, Y: D,

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Publications
[hide] Orphan missense mutations in the cystic fibrosis t... J Mol Diagn. 2011 Sep;13(5):520-7. Epub 2011 Jun 25. Fresquet F, Clement R, Norez C, Sterlin A, Melin P, Becq F, Kitzis A, Thoreau V, Bilan F
Orphan missense mutations in the cystic fibrosis transmembrane conductance regulator a three-step biological approach to establishing a correlation between genotype and phenotype.
J Mol Diagn. 2011 Sep;13(5):520-7. Epub 2011 Jun 25., [PMID:21708286]

Abstract [show]
More than 1860 mutations have been found within the human cystic fibrosis transmembrane conductance regulator (CFTR) gene sequence. These mutations can be classified according to their degree of severity in CF disease. Although the most common mutations are well characterized, few data are available for rare mutations. Thus, genetic counseling is particularly difficult when fetuses or patients with CF present these orphan variations. We describe a three-step in vitro assay that can evaluate rare missense CFTR mutation consequences to establish a correlation between genotype and phenotype. By using a green fluorescent protein-tagged CFTR construct, we expressed mutated proteins in COS-7 cells. CFTR trafficking was visualized by confocal microscopy, and the cellular localization of CFTR was determined using intracellular markers. We studied the CFTR maturation process using Western blot analysis and evaluated CFTR channel activity by automated iodide efflux assays. Of six rare mutations that we studied, five have been isolated in our laboratory. The cellular and functional impact that we observed in each case was compared with the clinical data concerning the patients in whom we encountered these mutations. In conclusion, we propose that performing this type of analysis for orphan CFTR missense mutations can improve CF genetic counseling.

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Sentences [show]
No. Sentence Comment
61 The genotype was as follows: p.[Lys696Arg] (K696R). Login to comment
97 Sequences of Site-Directed Mutagenesis Primers Mutation Sense oligonucleotide sequences L102P 5=-GTACAGCCTCTCTTACCGGGAAGAATCATAGCTTCC-3= L167R 5=-AAGAAGACTTTAAAGCGGTCAAGCCGTGTTCTAG-3= P574S 5=-GCTGATTTGTATTTATTAGACTCTTCTTTTGGATACCTAGATG-3= V562I 5=-AGAATTTCTTTAGCAAGAGCAATATACAAAGATGCTGATTTG-3= K696R 5=-CAGACTGGAGAGTTTGGGGAAAGAAGGAAGAATTCTATTCTC-3= P841R 5=-GATATGGAGAGCATACGAGCAGTGACTACATGG-3= CFTR Missense Mutation Biological Assay 3 JMD Month 2011, Vol. xx, No. Login to comment
109 Mixed Phenotype with P574S CFTR Variant The missense substitution p.[Pro574Ser] (P574S) lies within nucleotide-binding domain 1. Login to comment
119 Decreased Activity of K696R and P841R CFTR Mutants Two missense mutations, K696R and p.[Pro841Arg] (P841R), are situated within the cytoplasmic regulator domain of CFTR. Login to comment
123 [1210-12T(7)] ϩ [1210-12T(7)] CBAVD ϩ Ø Ø 5/F/37 p.[Lys696Arg] Asymptomatic (prenatal CF diagnostics) ϩ Ø Ø 6/M/33 c. Login to comment
133 After using Western blot analysis (Figure 3B), the proportions of core-glycosylated protein for K696R and P841R appear insignificantly different from that of WT-CFTR (n ϭ 3, data not shown). Login to comment
134 Moreover, we observe by iodide efflux experiments that both mutations have similar functional consequence on CFTR activity: the maximum activation is reduced to 49.67% Ϯ 2.61% (n ϭ 4) for K696R and to 45.10% Ϯ 4.94% (n ϭ 4) for P841R (Figure 3C). Login to comment
161 B A calreticulin egremS475P calreticulin egremS475P WT F508del P574S CFTR C WT F508del P574S CFTR C B CFTR NaKATPase B CFTR NaKATPase C WT P574S *** WT P574S ***** 00 WT P574S 0 00 00 00 00 **** 00 WT P574S 0 00 00 00 00 0 50 100 WT % of maximal activation 0 50 100 WT % of maximal activationB/(B+C) (%) 100 0 200 300 400 500 B/(B+C) (%) 100 0 200 300 400 500 Figure 2. Login to comment
162 P574S amino acid substitution decreases CFTR protein maturation. Login to comment
192 One possible explanation could be that iodide effluxes are not sensitive enough to precisely evaluate the CFTR chloride channel activation process; notably, CFTR channel open probability could not be studied directly by this A B C P841R actin merge +TO-PRO K696R actin merge +TO-PRO P841R actin merge +TO-PRO K696R actin merge +TO-PRO % of maximal activation ** ** 0 50 100 WT K696R P841R % of maximal activation ** ** 0 50 100 WT K696R P841R % of maximal activation ** ** 0 50 100 WT K696R P841R WT F508del P841R K696R B CFTR NaKATPase C WT F508del P841R K696R B CFTR NaKATPase C Figure 3. Login to comment
193 K696R and P841R amino acid substitutions affect CFTR channel activity. Login to comment
207 Obviously, patch-clamp analysis would be useful to measure the real impact of P574S in CFTR channel gating. Login to comment
209 Our results suggest a mild phenotype for the K696R and P841R CFTR mutants: the trafficking and maturation rates appear normal, whereas activation is approximately half reduced. Login to comment
211 The variant K696R was isolated from a familial study, in which we did not find any other CFTR mutation. Login to comment
60 Sequences of Site-Directed Mutagenesis Primers Mutation Sense oligonucleotide sequences L102P 5=-GTACAGCCTCTCTTACCGGGAAGAATCATAGCTTCC-3= L167R 5=-AAGAAGACTTTAAAGCGGTCAAGCCGTGTTCTAG-3= P574S 5=-GCTGATTTGTATTTATTAGACTCTTCTTTTGGATACCTAGATG-3= V562I 5=-AGAATTTCTTTAGCAAGAGCAATATACAAAGATGCTGATTTG-3= K696R 5=-CAGACTGGAGAGTTTGGGGAAAGAAGGAAGAATTCTATTCTC-3= P841R 5=-GATATGGAGAGCATACGAGCAGTGACTACATGG-3= was presented. Login to comment
62 The genotype was as follows: p.Lys696Arg (K696R). Login to comment
82 [1210-12T(7)] ϩ [1210-12T(7)] CBAVD ϩ Ø Ø 5/F/37 p.[Lys696Arg] Asymptomatic (prenatal CF diagnostics) ϩ Ø Ø 6/M/33 c. Login to comment
110 K696R and P841R amino acid substitutions affect CFTR channel activity. Login to comment
158 Decreased Activity of K696R and P841R CFTR Mutants Two missense mutations, p.Lys696Arg (K696R) and p.Pro841Arg (P841R), are situated within the cytoplasmic regulator domain of CFTR. Login to comment
205 Our results suggest a mild phenotype for the K696R and P841R CFTR mutants: the trafficking and maturation rates appear normal, whereas activation is approximately half reduced. Login to comment