ABCMdb

PMID: 12952861

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 [PubMed]

Sentences

No. Mutations Sentence Comment

4 ABCC7 p.Glu217Gly ABCC7 p.Gln1352His Several haplotypes, especially with Q1352H, IVS8 T5, and E217G, were found to have disease associations in a case-control study. Login to comment 7 ABCC7 p.Gln1352His In addition, a Q1352H mutation found in a V470 background showed the strongest disease association. Login to comment 9 ABCC7 p.Glu217Gly ABCC7 p.Gln1352His 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 10 ABCC7 p.Gln1352His Surprisingly, the additional M470V polymorphic variant with the Q1352H mutation completely abolished CFTR-dependent anion transport activities. Login to comment 15 ABCC7 p.Asn1303Lys ABCC7 p.Gly542* Several mutations of CFTR, such as F508del, G542X and N1303K, are associated with severe CF phenotypes and display high disease penetrance. Login to comment 44 ABCC7 p.Gln1352His In samples from patients, several other heteroduplexes were found including one in exon 22, which was identified as Q1352H (G to C change at nucleotide 4188). Login to comment 50 ABCC7 p.Arg117His Among the 10 worldwide disease-causing mutations, only one case of R117H was found in a control subject. Login to comment 52 ABCC7 p.Gln1352His Genetic variants at several sites were found to be associated with bronchiectasis and/or chronic pancreatitis, the most notable being Q1352H. Login to comment 53 ABCC7 p.Gln1352His The heterozygote frequency of Q1352H was significantly higher in bronchiectasis (P ¼ 0.02) and chronic pancreatitis patients (P ¼ 0.005) than in the control group. Login to comment 54 ABCC7 p.Glu217Gly 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 72 ABCC7 p.Gln1352His (C) The genetic variation in exon 22 was identified as Q1352H (4188G > C) by bi-directional nucleotide sequencing. Login to comment 74 ABCC7 p.Gly551Asp ABCC7 p.Arg117His ABCC7 p.Trp1282* ABCC7 p.Arg553* ABCC7 p.Asn1303Lys ABCC7 p.Gly542* ABCC7 p.Arg1162* ABCC7 p.Glu217Gly ABCC7 p.Arg1453Trp ABCC7 p.Ile556Val ABCC7 p.Ile125Thr ABCC7 p.Gln1352His 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 91 ABCC7 p.Gln1352His As expected from the genotype data, haplotype 4 containing Q1352H showed the strongest association with bronchiectasis and chronic pancreatitis in the Korean population (P ¼ 0.02 and P ¼ 0.008, respectively). Login to comment 92 ABCC7 p.Glu217Gly 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 ABCC7 p.Glu217Gly ABCC7 p.Gln1352His As shown in Figure 2A, the protein expressions of mature glycosylated CFTR were significantly decreased in E217G and Q1352H mutations. Login to comment 107 ABCC7 p.Arg117His ABCC7 p.Glu217Gly ABCC7 p.Arg1453Trp ABCC7 p.Ile556Val ABCC7 p.Ile125Thr ABCC7 p.Gln1352His 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 ABCC7 p.Arg117His ABCC7 p.Glu217Gly ABCC7 p.Arg1453Trp ABCC7 p.Ile556Val ABCC7 p.Ile125Thr ABCC7 p.Gln1352His 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 ABCC7 p.Glu217Gly ABCC7 p.Gln1352His 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 129 ABCC7 p.Arg1453Trp ABCC7 p.Gln1352His Among them, Q1352H showed the largest decrease (by 71%) and R1453W showed the smallest decrease (by 37%) in whole cell ClÀ currents (Fig. 3C). Login to comment 134 ABCC7 p.Arg1453Trp ABCC7 p.Ile556Val ABCC7 p.Gln1352His Compared with WT, Po was significantly reduced in I556V (by 34%), Q1352H (by 55%), and R1453W (by 78%). Login to comment 137 ABCC7 p.Glu217Gly ABCC7 p.Arg1453Trp ABCC7 p.Ile556Val 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 138 ABCC7 p.Gln1352His In the case of Q1352H, both membrane expression and Po were decreased. Login to comment 145 ABCC7 p.Glu217Gly ABCC7 p.Gln1352His 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 146 ABCC7 p.Gln1352His ABCC7 p.Gln1352His Effects of M470V on Q1352H The Q1352H mutation showed the highest disease associations in the genotype data. Login to comment 147 ABCC7 p.Gln1352His ABCC7 p.Gln1352His Since Q1352H is appeared only in the V470 background in the tested population (Table 3), we made M470V changes in WTand Q1352H-CFTR clones and performed functional studies. Login to comment 148 ABCC7 p.Gln1352His ABCC7 p.Gln1352His As shown in Figure 5A, additional M470V change did not alter the protein expression levels in either WTor Q1352H-CFTR. Login to comment 149 ABCC7 p.Gln1352His However, additional M470V change in Q1352H abolished cAMP-activated ClÀ currents (Fig. 5C and D) and cAMP-activated ClÀ /HCO3 À exchanges (Fig. 5E and F) by 99 and 95%, respectively. Login to comment 152 ABCC7 p.Arg117His Among the 10 common worldwide disease-causing mutations, only one case of R117H was found Figure 2. Login to comment 160 ABCC7 p.Asn1303Lys ABCC7 p.Gly542* However, over 95% of the three most common CF-causing mutations, F508del, G542X and N1303K, arise from this haplotype. Login to comment 176 ABCC7 p.Glu217Gly ABCC7 p.Arg1453Trp ABCC7 p.Ile556Val ABCC7 p.Gln1352His 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 186 ABCC7 p.Gln1352His Q1352H was found to be the most notable mutation in both case-control and molecular functional studies. Login to comment 191 ABCC7 p.Gln1352His However, its sequence is highly conserved in all vertebrates and the change of the last glutamine into histidine (Q1352H) evoked defects in protein Figure 4. Login to comment 196 ABCC7 p.Gln1352His Representative traces of Mock-, WTand Q1352H-transfected cells are shown in (A), and the results of five to six experiments for each mutant are summarized in (B). Login to comment 199 ABCC7 p.Gln1352His Effects of M470V on Q1352H mutant. Login to comment 200 ABCC7 p.Gln1352His M470V variation was introduced in the WT and the Q1352H clones and protein expressions and functional activities were measured using the protocols described in the legends of Figures 2-4. Login to comment 201 ABCC7 p.Gln1352His ABCC7 p.Gln1352His M470V change did not alter the protein expression levels in WTor Q1352H-CFTR (A, B). Login to comment 202 ABCC7 p.Gln1352His However, additional M470V change in Q1352H abolished cAMP-activated ClÀ currents (C, D) and ClÀ /HCO3 À exchanges (E, F). Login to comment 205 ABCC7 p.Glu217Gly 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 ABCC7 p.Glu217Gly These results imply that E217G is a mild mutation, which is also supported by our molecular data. Login to comment 207 ABCC7 p.Glu217Gly The E217G mutation partially decreased membrane protein expression and anion transporting activities by 60-70%. Login to comment 208 ABCC7 p.Glu217Gly ABCC7 p.Gln1352His 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 ABCC7 p.Glu217Gly ABCC7 p.Gln1352His 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 210 ABCC7 p.Gln1352His Therefore, combinatorial effects with M470V locus may influence the overall function of haploid gene products in Q1352H mutation. Login to comment 211 ABCC7 p.Gln1352His Indeed, molecular data of double mutagenesis showed a surprising result that the additional change of M470V in Q1352H abolished the anion transporting activities of CFTR protein (Fig. 5). Login to comment 213 ABCC7 p.Gln1352His ABCC7 p.Gln1352His A plausible scenario is that M470V and Q1352H might be involved in the cross talk and/or interaction between the two NBDs of CFTR (25,26), since M470V and Q1352H are located in the NBD1 and NBD2, respectively. Login to comment 226 ABCC7 p.Glu217Gly ABCC7 p.Gln1352His In conclusion, CFTR mutations of M470V-Q1352H, IVS8 T5-M470V, and E217G are associated with bronchiectasis and chronic pancreatitis. Login to comment 259 ABCC7 p.Glu217Gly ABCC7 p.Arg1453Trp ABCC7 p.Ile556Val ABCC7 p.Gln1352His 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