ABCMdb

PMID: 9736778

Vankeerberghen A, Wei L, Jaspers M, Cassiman JJ, Nilius B, Cuppens H
Characterization of 19 disease-associated missense mutations in the regulatory domain of the cystic fibrosis transmembrane conductance regulator.
Hum Mol Genet. 1998 Oct;7(11):1761-9., [PubMed]

Sentences

No. Mutations Sentence Comment

1 ABCC7 p.Ala613Thr ABCC7 p.Asp614Gly ABCC7 p.Gly628Arg ABCC7 p.Leu619Ser ABCC7 p.Ile618Thr ABCC7 p.His620Pro ABCC7 p.Leu610Ser ABCC7 p.Ile601Phe ABCC7 p.Leu633Pro Nine of these (I601F, L610S, A613T, D614G, I618T, L619S, H620P, G628R and L633P) resulted in aberrant processing. Login to comment 7 ABCC7 p.Gly622Asp ABCC7 p.Glu822Lys ABCC7 p.Arg792Gly Three mutant chloride channels, G622D, R792G and E822K CFTR, were characterized by significantly lower intrinsic chloride channel activities compared with wild-type CFTR. Login to comment 8 ABCC7 p.Ala800Gly ABCC7 p.His620Gln Two mutations, H620Q and A800G, resulted in increased intrinsic chloride transport activities. Login to comment 9 ABCC7 p.Glu826Lys ABCC7 p.Thr665Ser Finally, T665S and E826K CFTR had single channel properties not significantly different from wild-type CFTR. Login to comment 42 ABCC7 p.Arg792Gly ABCC7 p.Arg766Met ABCC7 p.Lys698Arg Of the 21 RD missense mutations, three (K698R, R766M and R792G) were located in PKA recognition consensus sites. Login to comment 43 ABCC7 p.Arg792Gly ABCC7 p.Arg766Met Only R766M and R792G disrupted a PKA recognition site and might therefore interfere with CFTR regulation. Login to comment 44 ABCC7 p.Val754Met ABCC7 p.Ala800Gly ABCC7 p.Asp648Val The different RD mutations affected almost all, except D648V, V754M and A800G, amino acid moieties that were absolutely or highly conserved. Login to comment 49 ABCC7 p.Phe693Leu ABCC7 p.Ile807Met Since then, it was found that the F693L mutation is a polymorphism (14,15) and that the I807M polymorphism is associated with congenital bilateral absence of the vas deferens (CBAVD) (our unpublished data). Login to comment 50 ABCC7 p.Val754Met ABCC7 p.Lys698Arg Two additional disease mutations, which were not characterized at the functional level in this study, were identified [K698R and V754M (15)]. Login to comment 60 ABCC7 p.Ile807Met This maturation pattern was found not only for wild-type CFTR but also for some of the mutants (Table 2), such as I807M CFTR (Fig. 2). Login to comment 61 ABCC7 p.Asp614Gly ABCC7 p.Leu610Ser Some of the mutants, however, presented an abnormal maturation pattern, as can be seen for L610S and D614G CFTR (Fig. 2). Login to comment 66 ABCC7 p.Ala613Thr ABCC7 p.Asp614Gly ABCC7 p.Gly628Arg ABCC7 p.Leu619Ser ABCC7 p.Ile618Thr ABCC7 p.His620Pro ABCC7 p.Leu610Ser ABCC7 p.Ile601Phe ABCC7 p.Leu633Pro The mutations that gave rise to a protein that was not able to proceed to the 190 kDa form (I601F, L610S, A613T, D614G, I618T, L619S, H620P, G628R and L633P; Table 2) are therefore class two mutations (17), where the disease phenotype is caused by the absence of sufficient CFTR protein at the cell surface. Login to comment 68 ABCC7 p.Ala613Thr ABCC7 p.Gly622Asp ABCC7 p.Glu826Lys ABCC7 p.Glu822Lys ABCC7 p.Arg792Gly ABCC7 p.Ala800Gly ABCC7 p.Arg766Met ABCC7 p.Phe693Leu ABCC7 p.Asp648Val ABCC7 p.His620Gln ABCC7 p.Asp614Gly ABCC7 p.Gly628Arg ABCC7 p.Thr665Ser ABCC7 p.Leu619Ser ABCC7 p.Ile618Thr ABCC7 p.Ile807Met ABCC7 p.His620Pro ABCC7 p.Leu610Ser ABCC7 p.Ile601Phe ABCC7 p.Leu633Pro Primers used for mutagenesis Primer Sequence I601F (a1933t) 5'-CTA ACA AAA CTA GGT TTT TGG TCA CTT C-3' L610S (t1961c) 5'-CTA AAA TGG AAC ATT CAA AGA AAG CTG-3' A613T (g1969a) 5'-CAT TTA AAG AAA ACT GAC AAA ATA TTA-3' D614G (a1973g) 5'-CAT TTA AAG AAA GCT GGC AAA ATA TTA A-3' I618T (t1985c) 5'-GAC AAA ATA TTA ACT TTG CAT GAA GG-3' L619S (t1988c) 5'-GAC AAA ATA TTA ATT TCG CAT GAA GGT-3' H620P (a1991c) 5'-CAA AAT ATT AAT TTT GCC TGA AGG TAG C-3' H620Q (t1992g) 5'-AAT ATT AAT TTT GCA GGA AGG TAG CAG-3' G622D (g1997a) 5'-TTG CAT GAA GAT AGC AGC TAT TTT TAT G-3' G628R (g2014c) 5'-GCA GCT ATT TTT ATC GGA CAT TTT C-3' L633P (t2030c) 5'-CAT TTT CAG AAC CCC AAA ATC TAC AGC-3' D648V (a2075t) 5'-CTC ATG GGA TGT GTT TCT TTC GAC C-3' T665S (a2125t) 5'-CAA TCC TAA CTG AGT CCT TAC ACC G-3' F693L (t2209c) 5'-CAG ACT GGA GAG CTT GGG GAA AAA AG-3' R766M (g2429t) 5'-GCA CGA AGG ATG CAG TCT GTC CTG-3' R792G (c2506g) 5'-CAG CAT CCA CAG GAA AAG TGT CAC TG-3' A800G (c2531g) 5'-CTG GCC CCT CAG GGA AAC TTG ACT G-3' I807M (a2553g) 5'-CTG AAC TGG ATA TGT ATT CAA GAA GG-3' E822K (g2596a) 5'-GGC TTG GAA ATA AGT AAA GAA ATT AAC G-3' E826K (g2608a) 5'-GAA GAA ATT AAC AAA GAA GAC TTA AAG-3' Selection primer BstBI 5'-CTC TGG GGT CCG GAA TGA CCG AC-3' Two primers were used for each mutagenesis reaction. Login to comment 77 ABCC7 p.Val754Met ABCC7 p.Ala613Thr ABCC7 p.Glu826Lys ABCC7 p.Glu822Lys ABCC7 p.Arg792Gly ABCC7 p.Ala800Gly ABCC7 p.Arg766Met ABCC7 p.Phe693Leu ABCC7 p.His620Gln ABCC7 p.Asp614Gly ABCC7 p.Gly628Arg ABCC7 p.Thr665Ser ABCC7 p.Leu619Ser ABCC7 p.Ile618Thr ABCC7 p.Ile807Met ABCC7 p.His620Pro ABCC7 p.Leu610Ser ABCC7 p.Ile601Phe ABCC7 p.Leu633Pro ABCC7 p.Lys698Arg Mutations detected in patients (I601F, L610S, A613T, D614G, I618T, L619S, H620P, H620Q, D622G, G628R, L633P, T665S, F693L, K698R, V754M, R766M, R792G, A800G, I807M, E822K and E826K) are indicated in bold and underlined, the PKA phosphorylation sites by an arrow and the two acidic domains are boxed. Login to comment 83 ABCC7 p.Glu826Lys ABCC7 p.Glu822Lys ABCC7 p.Arg792Gly ABCC7 p.Thr665Ser Four mutations (T665S, R792G, E822K and E826K) caused a significant reduction in the cAMP-induced chloride current. Login to comment 84 ABCC7 p.Ala800Gly ABCC7 p.His620Gln Interestingly, two mutations (H620Q and A800G) gave rise to chloride channels with significantly higher chloride transport activities. Login to comment 85 ABCC7 p.Gly622Asp ABCC7 p.Arg766Met ABCC7 p.Phe693Leu ABCC7 p.Asp648Val ABCC7 p.Ile807Met The remainder (G622D, D648V, F693L, R766M and I807M) did not significantly affect chloride transport ability when compared with wild-type CFTR channels. Login to comment 87 ABCC7 p.Trp1282* ABCC7 p.Gly542* ABCC7 p.Arg1158* ABCC7 p.Ala613Thr ABCC7 p.Gly622Asp ABCC7 p.Gly622Asp ABCC7 p.Glu826Lys ABCC7 p.Glu826Lys ABCC7 p.Glu822Lys ABCC7 p.Glu822Lys ABCC7 p.Arg792Gly ABCC7 p.Arg792Gly ABCC7 p.Arg792Gly ABCC7 p.Ala800Gly ABCC7 p.Ala800Gly ABCC7 p.Arg766Met ABCC7 p.Arg766Met ABCC7 p.Arg766Met ABCC7 p.Phe693Leu ABCC7 p.Phe693Leu ABCC7 p.Asp648Val ABCC7 p.Asp648Val ABCC7 p.His620Gln ABCC7 p.His620Gln ABCC7 p.Asp614Gly ABCC7 p.Asp614Gly ABCC7 p.Gly628Arg ABCC7 p.Thr665Ser ABCC7 p.Leu619Ser ABCC7 p.Leu619Ser ABCC7 p.Ile618Thr ABCC7 p.Ile618Thr ABCC7 p.Ile807Met ABCC7 p.Ile807Met ABCC7 p.His620Pro ABCC7 p.His620Pro ABCC7 p.Leu610Ser ABCC7 p.Ile601Phe ABCC7 p.Ile601Phe ABCC7 p.Leu633Pro ABCC7 p.Leu633Pro Maturation pattern of RD mutations and their associated phenotype found in patients with the indicated genotype (when the mutation is associated with CF, only the pancreas status is given) Mutation A-form B-form C-form Clinical data Genotype Phenotype Reference I601F + + - I601F/G542X PS M. Schwarz, personal communication L610S + + - Unknown Unknown A613T + + - Unknown Unknown D614G + + - D614G/unknown PI 14 I618T + + - I618T/dF508 PS G.R. Cutting, personal communication L619S + + - L619S/unknown PI B. Tümmler, personal communication H620P + + - H620P/R1158X PS M. Schwarz, personal communication H620Q + + + H620Q/dF508 PI T. Dörk, personal communication G622D + + + G622D/unknown Oligospermia J. Zielenski, personal communication G628R + + - Unknown Unknown L633P + + - L633P/3659delC M. Schwarz, personal communication D648V + + + D648V/3849+10kb C/T PI C. Ferec, personal communication T665S + + + Unknown Unknown F693L + + + F693L/W1282X Healthy C. Ferec; CF Genetic Analysis Consortium R766M + + + R766M/R792G CBAVD D. Glavac, personal communication R792G + + + R766M/R792G CBAVD D. Glavac, personal communication A800G + + + A800G/unknown CBAVD 34 I807M + + + I807M/unknown CBAVD Our observation E822K + + + E822K/unknown PI 35 E826K + + + E826K/unknown Thoracic sarcoidosis C. Bombieri, personal communication +, the protein matures up to that form; -, the protein does not reach the respective maturation step. Login to comment 97 ABCC7 p.Gly622Asp ABCC7 p.Glu822Lys ABCC7 p.Arg792Gly ABCC7 p.Ala800Gly ABCC7 p.His620Gln G622D, R792G and E822K gave rise to a CFTR chloride channel with a significantly lower Po than wild-type CFTR; H620Q and A800G CFTR resulted in channels with significantly higher Po. Login to comment 99 ABCC7 p.Asp614Gly ABCC7 p.Ile807Met ABCC7 p.Leu610Ser Pulse chase and immunoprecipitation of CFTR from COS1 cells transiently expressing wild-type, L610S, D614G or I807M CFTR. Login to comment 109 ABCC7 p.Ala613Thr ABCC7 p.Asp614Gly ABCC7 p.Gly628Arg ABCC7 p.Leu619Ser ABCC7 p.Ile618Thr ABCC7 p.His620Pro ABCC7 p.Leu610Ser ABCC7 p.Ile601Phe ABCC7 p.Leu633Pro Nine mutations caused aberrant processing: I601F, L610S, A613T, D614G, I618T, L619S, H620P, G628R and L633P. Login to comment 114 ABCC7 p.His620Gln When His620 was changed to Gln, noeffectonmaturationwasobserved.WhenHis620waschanged to Pro, an amino acid that dramatically affects the secondary structure of the protein and therefore possibly its correct folding, a non-maturing protein was obtained. Login to comment 123 ABCC7 p.Gly622Asp ABCC7 p.Glu826Lys ABCC7 p.Glu822Lys ABCC7 p.Arg792Gly ABCC7 p.Ala800Gly ABCC7 p.Arg766Met ABCC7 p.Phe693Leu ABCC7 p.Asp648Val ABCC7 p.His620Gln ABCC7 p.Thr665Ser ABCC7 p.Ile807Met Mutations that did not affect maturation (H620Q, G622D, D648V, T665S, F693L, R766M, R792G, A800G, I807M, E822K and E826K) were subsequently analysedat theelectrophysiologi- cal level. Login to comment 124 ABCC7 p.Gly622Asp ABCC7 p.Glu822Lys ABCC7 p.Arg792Gly Three of these (G622D, R792G and E822K) gave rise to chloride channels with significantly lower Po than the wild-type channel. Login to comment 125 ABCC7 p.Arg792Gly One of these mutations, R792G, disrupts a consensus recognition site for PKA. Login to comment 127 ABCC7 p.Ala800Gly ABCC7 p.His620Gln Strikingly, two mutations (H620Q and A800G) showed a significantly increased Po, when compared with wild-type CFTR. Login to comment 128 ABCC7 p.Pro574His So far, only the P574H mutation, located in NBD1, has been shown to give rise to a protein with higher intrinsic chloride transport properties. Login to comment 130 ABCC7 p.Ala800Gly ABCC7 p.His620Gln The 'hyperactive` disease-causing mutations described here, H620Q and A800G, do mature. Login to comment 131 ABCC7 p.Glu826Lys ABCC7 p.Arg766Met ABCC7 p.Phe693Leu ABCC7 p.Asp648Val ABCC7 p.Thr665Ser ABCC7 p.Ile807Met The remaining mutations (D648V, T665S, F693L, R766M, I807M and E826K) caused no significant alterations in intrinsic chloride channel activity. Login to comment 132 ABCC7 p.Phe693Leu F693L turned out to be a polymorphism (15), thereby explaining its normal function. Login to comment 136 ABCC7 p.His620Gln Activation of single channels by expressing wild-type and mutant CFTR (H620Q). Login to comment 137 ABCC7 p.His620Gln (A) Single channel current tracings of wild-type and H620Q CFTR before and after addition of the activation cocktail (1 µM IBMX and 0.1 µM forskolin) to the external site of the membrane patch. Login to comment