ABCMdb

PMID: 16429166

Brunham LR, Singaraja RR, Pape TD, Kejariwal A, Thomas PD, Hayden MR
Accurate prediction of the functional significance of single nucleotide polymorphisms and mutations in the ABCA1 gene.
PLoS Genet. 2005 Dec;1(6):e83. Epub 2005 Dec 30., [PubMed]

Sentences

No. Mutations Sentence Comment

38 ABCA1 p.Ser1731Cys All ABCA1 alleles expressed protein (R. Singaraja, H. Visscher, E. R. James, G. Chimini, and M. R. Hayden, unpublished data), with the exception of the S1731C cell line, for which we observed low levels of protein expression from two independently generated cell lines. Login to comment 39 ABCA1 p.Ser1731Cys ABCA1 p.Ser1731Cys To confirm that the S1731C allele was being expressed, we performed RT-PCR for ABCA1 on reverse-transcribed RNA from untransfected 293 cells and cells transfected with the wild-type or S1731C ABCA1 alleles. Login to comment 40 ABCA1 p.Ser1731Cys We found that cells transfected with the S1731C allele expressed abundant ABCA1 mRNA, at levels comparable to that of wild-type ABCA1 (Figure S1). Login to comment 41 ABCA1 p.Ser1731Cys The S1731C allele therefore expresses normal ABCA1 mRNA but fails to generate significant amounts of ABCA1 protein. Login to comment 45 ABCA1 p.Ile883Met ABCA1 p.Ile883Met ABCA1 p.Arg219Lys ABCA1 p.Arg219Lys ABCA1 p.Val771Met ABCA1 p.Val771Met ABCA1 p.Asp1289Asn ABCA1 p.Asp1289Asn ABCA1 p.Pro2150Leu ABCA1 p.Pro2150Leu À;3), R219K, V771M, I883M, D1289N, and P2150L, four had cholesterol efflux values that were not statistically different from wild-type ABCA1. Login to comment 46 ABCA1 p.Arg219Lys ABCA1 p.Val771Met ABCA1 p.Asp1289Asn ABCA1 p.Pro2150Leu This included two variants, D1289N and P2150L, that have been previously reported to be disease-causing mutations [4,8,9], as well as two cSNPs, R219K and V771M. Login to comment 47 ABCA1 p.Ile883Met One variant, I883M, was predicted to be functionally neutral but found to have cholesterol efflux modestly but significantly reduced (approximately 70% of wild-type ABCA1, p , 0.01). Login to comment 48 ABCA1 p.Trp590Ser ABCA1 p.Trp590Ser ABCA1 p.Ile883Met ABCA1 p.Ile883Met ABCA1 p.Arg219Lys ABCA1 p.Arg219Lys ABCA1 p.Gln597Arg ABCA1 p.Gln597Arg ABCA1 p.Cys1477Arg ABCA1 p.Cys1477Arg ABCA1 p.Asn1800His ABCA1 p.Asn1800His ABCA1 p.Ser1731Cys ABCA1 p.Ser1731Cys ABCA1 p.Lys776Asn ABCA1 p.Lys776Asn ABCA1 p.Val825Ile ABCA1 p.Val825Ile ABCA1 p.Met1091Thr ABCA1 p.Met1091Thr ABCA1 p.Thr929Ile ABCA1 p.Thr929Ile ABCA1 p.Arg587Trp ABCA1 p.Arg587Trp ABCA1 p.Asn935Ser ABCA1 p.Asn935Ser ABCA1 p.Arg230Cys ABCA1 p.Arg230Cys ABCA1 p.Val771Met ABCA1 p.Val771Met ABCA1 p.Ala937Val ABCA1 p.Ala937Val ABCA1 p.Glu1172Asp ABCA1 p.Glu1172Asp ABCA1 p.Val399Ala ABCA1 p.Val399Ala ABCA1 p.Arg1680Trp ABCA1 p.Arg1680Trp ABCA1 p.Asp1099Tyr ABCA1 p.Asp1099Tyr ABCA1 p.Phe2009Ser ABCA1 p.Phe2009Ser ABCA1 p.Ser1506Leu ABCA1 p.Ser1506Leu ABCA1 p.Leu1379Phe ABCA1 p.Leu1379Phe ABCA1 p.Thr774Pro ABCA1 p.Thr774Pro ABCA1 p.Asn935His ABCA1 p.Asn935His ABCA1 p.Asp1289Asn ABCA1 p.Asp1289Asn ABCA1 p.Ala255Thr ABCA1 p.Ala255Thr ABCA1 p.Ala1046Asp ABCA1 p.Ala1046Asp ABCA1 p.Arg1901Ser ABCA1 p.Arg1901Ser ABCA1 p.Val1704Asp ABCA1 p.Val1704Asp ABCA1 p.Pro2150Leu ABCA1 p.Pro2150Leu ABCA1 p.Pro85Leu ABCA1 p.Trp590Leu ABCA1 p.Trp590Leu ABCA1 p.Arg2081Trp ABCA1 p.Arg2081Trp ABCA1 p.Asn1611Asp ABCA1 p.Asn1611Asp ABCA1 p.Gln2196His ABCA1 p.Gln2196His ABCA1 p.Glu284Lys ABCA1 p.Glu284Lys ABCA1 p.Tyr482Cys ABCA1 p.Tyr482Cys ABCA1 p.His160Phe ABCA1 p.His160Phe This SNP has been reported to be associated with decreased HDL cholesterol and increased severity of atherosclerosis in Table 1. subPSEC Scores and Probability of Functional Impairment (Pdeleterious) for ABCA1 Mutations and SNPs Mutations SNPs Variant SubPSEC Pdeleterious Variant subPSEC Pdeleterious P85L À4.62 0.83 R219K À0.57 0.08 H160F À2.79 0.45 V399A À2.26 0.32 R230C À4.27 0.78 V771M À2.86 0.46 A255T À1.81 0.23 T774P À1.99 0.27 E284K À;2.34 0.34 K776N À;3.53 0.63 Y482C À4.21 0.77 V825I À1.06 0.13 R587W À6.04 0.95 I883M À1.38 0.17 W590S À5.19 0.9 E1172D À1.96 0.26 W590L À4.48 0.82 R1587K À0.58 0.08 Q597R À7.15 0.98 S1731C 0;4.21 0.77 T929I À4.29 0.78 N935H À8.54 1 N935S À7.53 0.99 A937V À6.6 0.97 A1046D 0;7.52 0.99 M1091T À3.56 0.64 D1099Y À6.09 0.96 D1289N À2.48 0.37 L1379F À3.81 0.69 C1477R À5.44 0.92 S1506L 0;5.17 0.9 N1611D À5.69 0.94 R1680W À6.02 0.95 V1704D À3.21 0.55 N1800H À4.23 0.77 R1901S À5.06 0.89 F2009S À2.73 0.43 R2081W À8.08 0.99 P2150L À2.88 0.47 Q2196H À2.74 0.43 DOI: 10.1371/journal.pgen.0010083.t001 PLoS Genetics | www.plosgenetics.org December 2005 | Volume 1 | Issue 6 | e83 0740 Accurate Prediction of ABCA1 Variants Synopsis A major goal of human genetics research is to understand how genetic variation leads to differences in the function of genes. Login to comment 58 ABCA1 p.Ser1731Cys The ABCA1 cSNP, S1731C, has a subPSEC score of less than À3, predictive of a deleterious effect on ABCA1 function. Login to comment 59 ABCA1 p.Ser1731Cys Cells transfected with the S1731C allele displayed a significant reduction in cholesterol efflux, relative to wild-type ABCA1 (p , 0.01), indicating that this SNP significantly impairs ABCA1 function, as predicted by PANTHER. Login to comment 60 ABCA1 p.Ser1731Cys These data indicate that S1731C may be a useful SNP to use as a functional marker in association studies. Login to comment 61 ABCA1 p.Met1091Thr Of all ABCA1 alleles tested functionally, M1091T displays the greatest reduction in cholesterol efflux (6.9 6 20% of wild-type ABCA1), consistent with previous reports that this is a severe mutation associated with a severe clinical presentation [4,13]. Login to comment 63 ABCA1 p.Met1091Thr ABCA1 p.Met1091Val ABCA1 p.Met1091Leu To determine whether the severe phenotype conferred by the M1091T mutation is a result of the sensitivity of this site, or rather is specific to the insertion of the threonine residue, we generated and characterized cell lines transfected with plasmids bearing M1091L and M1091V alleles, both predicted to have no impact on ABCA1 function (subPSEC scores À2.65 and À2.71, respectively). Login to comment 64 ABCA1 p.Met1091Thr Interestingly, both of these mutations dramatically impair cholesterol efflux, to a similar extent as the M1091T mutation (Figure 2B). Login to comment 75 ABCA1 p.Trp590Ser ABCA1 p.Trp590Ser ABCA1 p.Ile883Met ABCA1 p.Ile883Met ABCA1 p.Arg219Lys ABCA1 p.Arg219Lys ABCA1 p.Gln597Arg ABCA1 p.Gln597Arg ABCA1 p.Cys1477Arg ABCA1 p.Cys1477Arg ABCA1 p.Asn1800His ABCA1 p.Asn1800His ABCA1 p.Ser1731Cys ABCA1 p.Ser1731Cys ABCA1 p.Met1091Thr ABCA1 p.Met1091Thr ABCA1 p.Thr929Ile ABCA1 p.Thr929Ile ABCA1 p.Arg587Trp ABCA1 p.Arg587Trp ABCA1 p.Asn935Ser ABCA1 p.Asn935Ser ABCA1 p.Val771Met ABCA1 p.Val771Met ABCA1 p.Ser1506Leu ABCA1 p.Ser1506Leu ABCA1 p.Asp1289Asn ABCA1 p.Asp1289Asn ABCA1 p.Ala1046Asp ABCA1 p.Ala1046Asp ABCA1 p.Pro2150Leu ABCA1 p.Pro2150Leu ABCA1 p.Arg2081Trp Cholesterol Efflux Values for 293 Cells Transfected with ABCA1 Variants and subPSEC and PolyPhen Predictions of the Functional Impact of these Variants Variant Variant Type subPSEC Cholesterol Efflux PolyPhen R2081W Mutation À8.08 21.1 6 21%* Probably damaging N935S Mutation À7.53 29.3 6 13%* Benign A1046D Mutation À7.52 16.8 6 7%* Possibly damaging Q597R Mutation À7.15 17.7 6 14%* Probably damaging R587W Mutation À6.04 31.7 6 33%* Probably damaging C1477R Mutation À5.44 20.5 6 10%* Probably damaging W590S Mutation À5.19 47.1 6 13%* Probably damaging S1506L Mutation À5.17 17.8 6 15%* Probably damaging T929I Mutation À4.29 69.9 6 11%* Possibly damaging N1800H Mutation À4.23 31.3 6 16%* Possibly damaging S1731C SNP À4.21 12.3 6 10%* Possibly damaging M1091T Mutation À3.56 6.9 6 20%* Probably damaging P2150L Mutation À2.88 88.4 6 21% Probably damaging V771M SNP À2.86 145.4 6 33% Benign D1289N Mutation À2.48 137.7 6 86% Benign I883M SNP À1.38 69.1 6 16%* Benign R219K SNP À0.57 103.7 6 21.05 Benign Wild-type - 0.0 100% - *p , 0.01 compared to wild-type ABCA1. Login to comment 87 ABCA1 p.Met1091Thr ABCA1 p.Met1091Val ABCA1 p.Met1091Leu (B) Cholesterol efflux was assessed in 293 cells stably transfected with wild-type, M1091T, M1091L, or M1091V ABCA1 alleles. Login to comment 93 ABCA1 p.Asn935Ser ABCA1 p.Pro2150Leu harvard.edu/pph) is a Web-based program used to predict allele function based on homology and three-dimensional structural models where available [15], and it predicts alleles as being ''probably damaging,`` ''possibly damaging,`` or ''benign.`` In the dataset of variants for which we assessed cholesterol efflux, the predictions made by PANTHER and PolyPhen were significantly different for two mutations: N935S and P2150L. Login to comment 94 ABCA1 p.Asn935Ser PolyPhen predicted N935S to be benign, while PANTHER predicted it to be deleterious. Login to comment 95 ABCA1 p.Pro2150Leu Conversely, PolyPhen predicted P2150L to be probably damaging, while PANTHER predicted it to be neutral. Login to comment 97 ABCA1 p.Leu1026Pro Using the set of 12 ABCA1 variants described by Cohen et al. [14] for which macrophage efflux is reported as a second dataset, the PANTHER prediction differs significantly from PolyPhen in one case, where PANTHER correctly predicts that L1026P will be deleterious (efflux rates 2 SDs below control levels), while PolyPhen predicts the substitution will be benign. Login to comment 106 ABCA1 p.Asp1289Asn ABCA1 p.Pro2150Leu Both the D1289N and P2150L mutations are reported as pathogenic and causative of disease in the TD patients in which they were identified [4,8,9]. Login to comment 110 ABCA1 p.Trp590Ser ABCA1 p.Trp590Ser ABCA1 p.Asn1800His ABCA1 p.Asn1800His ABCA1 p.Ser1731Cys ABCA1 p.Ser1731Cys ABCA1 p.Ser1181Phe ABCA1 p.Ser1181Phe ABCA1 p.Cys1477Phe ABCA1 p.Cys1477Phe ABCA1 p.Leu1026Pro ABCA1 p.Leu1026Pro ABCA1 p.Glu1386Gln ABCA1 p.Glu1386Gln ABCA1 p.Arg965Cys ABCA1 p.Arg965Cys ABCA1 p.Asp1706Asn ABCA1 p.Pro85Leu ABCA1 p.Pro85Leu ABCA1 p.His551Asp ABCA1 p.His551Asp ABCA1 p.Trp590Leu ABCA1 p.Trp590Leu ABCA1 p.Pro248Ala ABCA1 p.Pro248Ala ABCA1 p.Ala1670Thr ABCA1 p.Ala1670Thr ABCA1 p.Thr2073Ala ABCA1 p.Thr2073Ala ABCA1 p.Arg306His ABCA1 p.Arg306His ABCA1 p.Glu815Gly ABCA1 p.Glu815Gly ABCA1 p.Arg638Gln ABCA1 p.Arg638Gln ABCA1 p.Arg1341Thr ABCA1 p.Arg1341Thr ABCA1 p.Arg1615Gln ABCA1 p.Arg1615Gln ABCA1 p.Ser1376Gly ABCA1 p.Ser1376Gly ABCA1 p.Thr459Pro ABCA1 p.Thr459Pro ABCA1 p.Asp2243Glu ABCA1 p.Asp2243Glu ABCA1 p.Lys401Gln ABCA1 p.Lys401Gln DOI: 10.1371/journal.pgen.0010083.g003 Table 3. subPSEC Scores for ABCA1 Variants Described in a Cohort of Individuals with Low HDL Cholesterol from the General Population Variant subPSEC Score Macrophage Efflux PolyPhen D1706N À6.57 0.38a Possibly damaging C1477F À5.55 0.34a Probably damaging W590S À5.19 - Probably damaging H551D À4.99 0.32a Probably damaging P85L À4.62 0.8 Probably damaging W590L À;4.48 0.31a Probably damaging N1800H À;4.23 0.27a Possibly damaging R965C À4.22 0.59 Probably damaging S1731C À4.21 0.28a Possibly damaging A1670T À;4.2 - Possibly damaging K401Q À;4.2 - Benign T459P À4.11 0.28a Possibly damaging R638Q À4.08 - Possibly damaging L1026P À3.86 0.25a Benign T2073A À3.84 0.28a Possibly damaging E815G À3.53 - Probably damaging R1615Q À3.45 - Possibly damaging S1181F À3.44 - Possibly damaging R306H À;3.31 - Benign E1386Q 0;2.44 0.51 Benign S1376G À2.19 - Benign R1341T À2.09 - Possibly damaging D2243E À1.6 - Benign P248A À0.18 - Benign a Efflux value is 2 SDs or more below control levels of 0.52 6 0.07. Login to comment 114 ABCA1 p.Asp1289Asn ABCA1 p.Asp1289Asn ABCA1 p.Arg2081Trp The TD patient described with the D1289N variant was also homozygous for a second mutation, R2081W [9], and our results strongly suggest that it is this second mutation, and not D1289N, that causes the phenotype observed in that patient. Login to comment 115 ABCA1 p.Pro2150Leu The molecular cause of the phenotype in patients carrying the P2150L variant remains to be determined, and it is possible that these patients harbor a second, yet unidentified coding or noncoding variant. Login to comment 119 ABCA1 p.Asp1289Asn ABCA1 p.Pro2150Leu Our efflux data showing that the D1289N and P2150L mutations are functionally neutral confirm the prediction that the conservation of these residues among ABCA1 proteins is not due to functional constraint, but rather reflects their recent common ancestry. Login to comment 120 ABCA1 p.Ser1731Cys One ABCA1 cSNP, S1731C, had a subPSEC score less than À3. Login to comment 122 ABCA1 p.Ser1731Cys Individuals carrying both the 2144X mutation and S1731C had significantly lower HDL cholesterol than individuals with only the 2144X mutation, although the number of patients in each group was small [10]. Login to comment 123 ABCA1 p.Ser1731Cys Cells transfected with the S1731C allele expressed ABCA1 mRNA at levels comparable to wild-type ABCA1; however, this cell line expressed low levels of ABCA1 protein and was markedly deficient in cholesterol efflux. Login to comment 126 ABCA1 p.Ser1731Cys The identification of S1731C as a functionally significant variant indicates that it may be a useful DNA marker to be used in association studies. Login to comment 138 ABCA1 p.Met1091Thr The subPSEC score for the naturally occurring M1091T mutation (À3.56) is only marginally predictive of a negative impact on function, but this variant resulted in a severe reduction in ABCA1 function, consistent with the severe phenotype observed in patients harboring this mutation [4,13]. Login to comment 139 ABCA1 p.Met1091Val ABCA1 p.Met1091Leu In addition, both the M1091V and M1091L substitutions severely impaired the function of ABCA1, yet they were predicted to be functionally neutral. Login to comment 142 ABCA1 p.Met1091Thr ABCA1 p.Met1091Val ABCA1 p.Met1091Leu Therefore, when calculating amino acid probabilities for position 1091, the subPSEC method includes sequences from only ABCA1 and ABCA4, which represents enough sequence variability to predict that a relatively radical mutation such as M1091T will likely be deleterious but not enough to predict that relatively conservative mutations such as M1091L or M1091V will be deleterious. Login to comment 143 ABCA1 p.Met1091Leu Our experimental finding that M1091L severely impairs ABCA1 cholesterol efflux suggests that substitution of leucine for methionine at this position may have played an important role in the functional divergence of ABCA7 from ABCA1. Login to comment 147 ABCA1 p.Ile883Met The I883M substitution results in a milder phenotype, with a modest but significant reduction in ABCA1-mediated cholesterol efflux. Login to comment 150 ABCA1 p.Ile883Met This divergence explains why a simple conservation-based approach predicts that I883M is a neutral substitution. Login to comment 238 ABCA1 p.Ser1731Cys ABCA1 p.Ser1731Cys Expression of the S1731C Allele in Polyclonal Stable Cell Lines ABCA1 protein (A) and mRNA (B) expression levels were determined in an untransfected control cell line, and cells transfected with wild-type ABCA1 or the S1731C variant. Login to comment 239 ABCA1 p.Ser1731Cys The cell line transfected with the S1731C allele expressed low levels of protein (A), but normal levels of mRNA (B), indicating that this variant impairs ABCA1 function by inhibiting the generation of a stable protein. Login to comment