ABCMdb

PMID: 12394343

Rohlfs EM, Zhou Z, Sugarman EA, Heim RA, Pace RG, Knowles MR, Silverman LM, Allitto BA
The I148T CFTR allele occurs on multiple haplotypes: a complex allele is associated with cystic fibrosis.
Genet Med. 2002 Sep-Oct;4(5):319-23., [PubMed]

Sentences

No. Mutations Sentence Comment

0 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr The I148T CFTR allele occurs on multiple haplotypes: A complex allele is associated with cystic fibrosis Elizabeth M. Rohlfs, PhD1 , Zhaoqing Zhou, PhD1 , Elaine A. Sugarman, MS1 , Ruth A. Heim, PhD1 , Rhonda G. Pace, BS2 , Michael R. Knowles, MD2 , Lawrence M. Silverman, PhD3 , and Bernice A. Allitto, PhD1 Purpose: To determine whether CFTR intragenic changes modulate the cystic fibrosis (CF) phenotype in individuals who are positive for the I148T allele. Login to comment 1 ABCC7 p.Ile148Thr Methods: The CFTR genes from individuals who carried at least one copy of the I148T allele were analyzed for additional changes that may be acting as genetic modifiers. Login to comment 2 ABCC7 p.Ile148Thr Results: Seven of eight individuals with a known or suspected diagnosis of CF who carried I148T in combination with a severe CF mutation also carried 3199del6. Login to comment 3 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr Eight apparently healthy adult individuals who were compound heterozygous for I148T and a severe CF mutation or homozygous for I148T did not carry the deletion (P ϭ 0.0014). Login to comment 4 ABCC7 p.Ile148Thr The I148T allele occurs on at least three haplotypes: an IVS-8 9T background, a 7T background, or a 9T ϩ 3199del6 background. Login to comment 6 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr Conclusions: It is concluded that I148T occurs on at least three haplotypes and the complex allele I148T ϩ 9T ϩ 3199del6 is associated with a classic CF phenotype. Login to comment 12 ABCC7 p.Arg117His ABCC7 p.Arg117His ABCC7 p.Arg117His Most notable is the effect of the length of the intron 8 polythymidine tract (5, 7, or 9 thymidines) on exon 9 splicing and the phenotypic expression of the R117H mutation.7,8 Individuals with a severe CF mutation and R117H in cis with 5 thymidines (5T) may have moderate (pancreatic sufficient) CF, whereas those with R117H and 7 thymidines (7T) in cis may be asymptomatic and have CBAVD or later-onset lung disease such as bronchiectasis.8,9 Only a small number of all CFTR mutations result in a predict- ablediseasephenotype.Theclassificationisusuallybasedoncare- ful clinical description of patients with defined genotypes. Login to comment 13 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr The I148T allele has been identified in CF patients from several countries and has been characterized as a "severe" CF mutation be- causesomeofthesepatientshadpancreaticinsufficientCF.10-13 It accounts for 0.7% of CF chromosomes in Italian patients,12 0.2% in Canadian patients,14 1.1% in US patients from the state of Maine,15 0.08% in French patients,16 and 9.1% in French-Canadian patients.17 In addition, the substitution changes a conserved amino acid and occurs in the first cytoplasmic loop of the first membrane spanning domain (MSD1).18,19 Based on such data, I148T was included in the ACMG/ACOG recommended CF mutation screening panel.20,21 Functional analysis of the I148T allele in transfected cell lines indicates that the protein product from this allele is fully glycosylated and biosynthetically mature. Login to comment 14 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr Anion conduction, as measured by iodide efflux, is normal in cells containing the I148T allele, and the cells have gating properties identical with those of wild-type cells.19 While this analysis does not rule out the possibility that the I148T allele affects other functions, such as its ability to function as a sodium channel, several major CFTR functions are retained. Login to comment 21 ABCC7 p.Ile148Thr 5 a r t i c l e Genetics IN Medicine We now have evidence that the I148T allele is associated with variable phenotypes. Login to comment 22 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr First, we had previously identified apparently healthy individuals who were compound heterozygotes for ⌬F508 and I148T.22 Second, we observed that the frequency of I148T is significantly higher in CF carriers than CF patients.22,23 To determine whether additional changes within the CFTR gene were influencing the CF phenotype when I148T was present, we sequenced the coding regions of the CFTR gene in apparently healthy individuals and CF patients who were compound heterozygous or homozygous for I148T. Login to comment 23 ABCC7 p.Ile148Thr Here we show that the variable phenotypes are due to the occurrence of an in-frame deletion in cis with I148T on some chromosomes. Login to comment 25 ABCC7 p.Ile148Thr No age limit was applied when determining the frequency of I148T in carriers, CF patients, or suspected CF patients. Login to comment 27 ABCC7 p.Ile148Thr As described below, carriers, CF patients, and suspected CF patients who carried I148T and a severe mutation were tested for the 3199del6 allele. Login to comment 28 ABCC7 p.Ile148Thr Carriers of a single I148T allele were also tested for 3199del6. Login to comment 29 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr Individuals homozygous for I148T or compound heterozygous for I148T and a severe CF allele were ascertained from January 1, 1998, to December 1, 2001. Login to comment 38 ABCC7 p.Ile148Thr Sequence analysis To search for additional sequence changes in individuals who carried the I148T allele, coding exons of the CFTR gene were sequenced by single PCR amplifications spanning each exon followed by BigDye terminator cycle sequencing. An ABI 310 automated sequencer was used to analyze the cycle sequencing products. Login to comment 49 ABCC7 p.Ile148Thr RESULTS Frequency of I148T in carriers, CF patients, and suspected CF patients Carrier testing of 42,784 individuals without a family history of CF identified 1,754 individuals who carried one mutation (carrier frequency approximately 1/24). Login to comment 50 ABCC7 p.Ile148Thr One hundred thirteen of the identified mutations were I148T (6.4%). Login to comment 52 ABCC7 p.Ile148Thr However, only 6 (0.06%) of the affected CF chromosomes were I148T. Login to comment 53 ABCC7 p.Ile148Thr This represents a 106-fold difference in frequency of I148T between the carrier and affected groups. Login to comment 54 ABCC7 p.Ile148Thr In a group of 10,231 patients referred for testing due to a suspected diagnosis of CF, 1.1% of the identified CF chromosomes were I148T. Login to comment 55 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr CFTR analysis of apparently healthy individuals and CF patients with the I148T allele Carrier testing identified seven compound heterozygotes (⌬F508/I148T) and one I148T homozygote who were apparently unaffected with CF (Table 1). Login to comment 57 ABCC7 p.Ile148Thr The presence of the I148T and ⌬F508 on the same chromosome was ruled out in two individuals by genotyping of additional family members. Login to comment 58 ABCC7 p.Asn1303Lys ABCC7 p.Ile148Thr ABCC7 p.Gln890* Diagnostic testing of individuals affected with CF, or suspected of having CF, identified eight individuals who were compound heterozygous for a severe mutation (⌬F508, N1303K, or Q890X) and I148T (Table 2). Login to comment 61 ABCC7 p.Ile148Thr Analysis of the intron 8 polythymidine tract determined that there was no significant difference in the poly(T) background of the I148T chromosomes between the healthy and affected groups (Tables 1 and 2). Login to comment 62 ABCC7 p.Ile148Thr The I148T allele appears to occur most frequently on a 9T chromosomal background. Login to comment 65 ABCC7 p.Ile148Thr However, in individual 38 it is most likely that the ⌬F508 is on the 9T chromosome and the I148T is on the 7T chromosome.7 Analysis of additional CFTR exons from healthy and affected patients identified a 6-bp deletion at nucleotide 3199 in some individuals. Login to comment 73 ABCC7 p.Ile148Thr Sequencing of all exons did not identify any CFTR mutations in addition to I148T and ⌬F508. Login to comment 74 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr None of the eight apparently healthy individuals (⌬F508/I148T or I148T/ I148T) carried the deletion. Login to comment 75 ABCC7 p.Ile148Thr The frequency of 3199del6 in individuals who are compound heterozygous or homozygous for I148T and not affected with CF versus those who are affected with or suspected of having CF is significantly different (P ϭ 0.0014 by Fisher exact test). Login to comment 76 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr Analysis of 3199del6 on other chromosomal backgrounds Inasmuch as one individual in the healthy group is presumed to carry I148T on a 7T chromosomal background, we Table 1 3199del6 analysis in apparently healthy adult individuals compound heterozygous or homozygous for I148T Indiv no. Indication for testing Ethnicity Age (yr) Mutation 1 Mutation 2 Intron 8 poly(T) 3199del6 12 Carrier testing w/FH NEC 56 I148T I148T 9, 9 neg 13 Screening NEC 40 ⌬F508 I148T 9, 9 neg 14 Carrier testing w/FH Hispanic 32 ⌬F508 I148T 9, 9 neg 15 Carrier testing w/FH NEC 29 ⌬F508 I148T 9, 9 neg 16 Carrier testing w/FH NEC 21 ⌬F508 I148T 9, 9 neg 23 Screening NEC 34 ⌬F508 I148T 9, 9 neg 38 Carrier testing w/FH NEC, SEC 31 ⌬F508 I148T 7, 9 neg 40 Carrier testing Caucasian 30 ⌬F508 I148T 9, 9 neg FH, family history; NEC, Northern European Caucasian; SEC, Southern European Caucasian. Login to comment 77 ABCC7 p.Asn1303Lys ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Gln890* Table 2 3199del6 analysis in patients with a known or suspected diagnosis of CF and compound heterozygous for I148T and a severe CF mutation Indiv no. Indication for testing Ethnicity Age (yr) Mutation 1 Mutation 2 Intron 8 poly(T) 3199del6 17 Suspected CF NEC 4 ⌬F508 I148T 9, 9 pos 18 Suspected CF Caucasian, N. American 1 ⌬F508 I148T 9, 9 pos 19 Affected CF NEC Ͻ1 N1303K I148T 9, 9 pos 20 Suspected CF NEC 5 ⌬F508 I148T 9, 9 nega 22 Affected CF NEC 14 ⌬F508 I148T 9, 9 pos 39 Pancreatic steatorrhea NEC 3 ⌬F508 I148T 9, 9 pos 41 Suspected CF Hispanic 10 Q890X I148T 7, 9 pos 42 Affected CF NEC 20 ⌬F508 I148T 9, 9 pos NEC, Northern European Caucasian. Login to comment 79 ABCC7 p.Ile148Thr tested 14 individuals who carried one copy of I148T, and who were homozygous for 7T, for the 3199del6 deletion (Table 3). Login to comment 80 ABCC7 p.Ile148Thr From this analysis, the I148T-7T haplotype does not appear to carry the 3199del6 deletion. Login to comment 81 ABCC7 p.Ile148Thr Poly(T) analysis of 76 additional individuals who carried one copy of I148T identified 8 who were homozygous for 9T and 64 who were compound heterozygous for 7T and 9T. Login to comment 83 ABCC7 p.Ile148Thr Of those individuals whose indication for testing was screening, 1 of 55 I148T carriers also carried the deletion. Login to comment 84 ABCC7 p.Ile148Thr Since we did not identify any individuals who were 5T/5T, we were unable to determine whether I148T occurs on a 5T background. Login to comment 85 ABCC7 p.Ile148Thr To determine whether 3199del6 occurred on CF chromosomes without I148T, we analyzed 95 CF patients with one or no identified CFTR mutations for the deletion. Login to comment 90 ABCC7 p.Arg117His ABCC7 p.Ile148Thr This was first appreciated when the frequency of R117H in carriers was observed to be 16-fold higher than the frequency in CF patients.25 In the ethnically diverse US population reported here, the I148T allele is 100-fold more frequent in carriers than in CF patients. Login to comment 92 ABCC7 p.Arg117His ABCC7 p.Arg117His There are now several published reports of ⌬F508/R117H compound heterozygotes that are apparently healthy.22,25,26 The R117H mutation was first identified in 199027 and has been included in most routine CF genotyping programs, increasing the likelihood that these individuals would have been detected as large numbers of individuals were tested. Login to comment 93 ABCC7 p.Ile148Thr In contrast, while considered a severe mutation, I148T has not been included in most routine testing. Login to comment 94 ABCC7 p.Ile148Thr This has likely limited the detection of healthy carriers of I148T/⌬F508 up to this time. Login to comment 95 ABCC7 p.Ile148Thr In addition, screening of affected CF populations by allele-specific assays continues to identify I148T in those with CF. Login to comment 97 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr The 3199del6 mutation was reported to the CF Consortium Mutation Database in 1998, and it was noted to occur on the same chromosome as I148T.28 Our data corroborate this finding since the CF patients with the deletion and I148T also carried one of three different severe CF alleles. Login to comment 105 ABCC7 p.Ile148Thr The deletion is apparently a rare allele, inasmuch as it was identified in only 2 of 90 individuals who carried one copy of I148T and was not identified in 386 non-CF chromosomes. Login to comment 106 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr It appears most likely that 3199del6 arose on a chromosome that carried the I148T mutation, and thus all chromosomes that have the deletion also have I148T. Login to comment 108 ABCC7 p.Asp1270Asn ABCC7 p.Arg74Trp For example, the complex allele composed of R74W and D1270N is known to produce variable phenotypes. Login to comment 109 ABCC7 p.Arg74Trp ABCC7 p.Ile148Thr In vitro studies of chloride channel function showed that R74W Table 3 3199del6 and poly(T) analysis in individuals who carry one copy of I148T Intron 8 poly (T) Indication for testing no. Login to comment 111 ABCC7 p.Asp1270Asn ABCC7 p.Asp1270Asn ABCC7 p.Arg74Trp has little deleterious effect and could be considered a polymorphism.31 D1270N has a more substantial effect on chloride channel function in this model, with the greatest effect observed in the cells containing the complex allele of R74W ϩ D1270N. Login to comment 112 ABCC7 p.Asp1270Asn ABCC7 p.Arg74Trp When in combination with a severe allele on the other chromosome, R74W ϩ D1270N is disease-causing; it was identified in a male with CBAVD, rhinitis, and recurrent respiratory infections.32 However,itwasalsodetectedina25-year-oldwomanwhoseonly clinicalsymptomofCFweretwopositivesweatchloridetests.33 In addition, genotyping programs that detect alleles of unknown functionalconsequence,andunknownpopulationfrequency,are at risk for erroneously reporting two changes in trans that are in fact a complex allele. Login to comment 113 ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr ABCC7 p.Ile148Thr WehavenowdeterminedthattheI148Talleleoccursonatleast three chromosomal backgrounds (I148T ϩ 7T, I148T ϩ 9T, and I148T ϩ 9T ϩ 3199del6) and that only the complex allele, I148T ϩ 9T ϩ 3199del6, appears to be associated with a classic CF phenotype. Login to comment 114 ABCC7 p.Ile148Thr Our data do not support the occurrence of the deletion alone;therefore,wecannotassesswhetherthecombinationofthe missense mutation and the deletion on the same chromosome is pathogenic or whether the entire phenotypic effect is due to the deletion.Withthecurrentrecommended25mutationCFscreen- ing panel in place,20,21 we recommend that an I148T positive result be followed by reflex testing for the 3199del6 mutation until the data presented here and from other laboratories are reviewed by the ACMG/ACOG Committee. Login to comment 115 ABCC7 p.Ile148Thr The presence of the complex allele in combination with another severe CF allele confers a CF phenotype, whereas I148T alone, even when a severe allele is on the other chromosome, confers no apparent phenotype. Login to comment 116 ABCC7 p.Ile148Thr Long-term follow-up studies may be necessary to confidently rule out an association with I148T alone and single-organ or late-onset clinical expression. Login to comment