Disease phenotypes and defects in function can be traced to nonsynonymous single nucleotide polymorphisms (nsSNPs), which are important indicators of action sites and effective potential therapeutic approaches. Identification of deleterious nsSNPs is crucial to characterize the genetic basis of diseases, assess individual susceptibility to disease, determinate molecular and therapeutic targets, and predict clinical phenotypes. In this study using PolyPhen2 and MutPred in silico algorithms, we analyzed the genetic variations that can alter the expression and function of the ABCA1 gene that causes the allelic disorders familial hypoalphalipoproteinemia and Tangier disease. Predictions were validated with published results from in vitro, in vivo, and human studies. Out of a total of 233 nsSNPs, 80 (34.33%) were found deleterious by both methods. Among these 80 deleterious nsSNPs found, 29 (12.44%) rare variants resulted highly deleterious with a probability >0.8. We have observed that mostly variants with verified functional effect in experimental studies are correctly predicted as damage variants by MutPred and PolyPhen2 tools. Still, the controversial results of experimental approaches correspond to nsSNPs predicted as neutral by both methods, or contradictory predictions are obtained for them. A total of seventeen nsSNPs were predicted as deleterious by PolyPhen2, which resulted neutral by MutPred. Otherwise, forty two nsSNPs were predicted as deleterious by MutPred, which resulted neutral by PolyPhen2. 1. Introduction Nonsynonymous single nucleotide polymorphisms (nsSNPs) are single base changes in coding regions that cause an amino acid substitution in the correspondent proteins. These missense variants constitute the most identifiable group of SNPs represented by a small (<1%) proportion . The nsSNPs might alter structure, stability, and function of proteins and produce the least conservative substitutions with drastic phenotypic consequences [2–5]. Studies suggest that about 60% of Mendelian diseases are caused by amino acid exchanges . Thousands of associations between Mendelian and complex diseases reveal a phenotypic code that links each complex disorder to a unique set of Mendelian loci . Discriminating disease-associated from neutral variants would help to understand the genotype/phenotype relation and to develop diagnosis and treatment strategies for diseases. Nonetheless, the most important application is the evaluation of functional effect and impact of genomic variation, relating interactions with phenotypes translating the finding
C. M. Yates and M. J. E. Sternberg, “The effects of non-synonymous single nucleotide polymorphisms (nsSNPs) on protein-protein interactions,” Journal of Molecular Biology, vol. 425, no. 21, pp. 3949–3969, 2013.
M. González-Castejón, F. Marín, C. Soler-Rivas, G. Reglero, F. Visioli, and A. Rodríguez-Casado, “Functional non-synonymous polymorphisms prediction methods: current approaches and future developments,” Current Medicinal Chemistry, vol. 18, no. 33, pp. 5095–5103, 2011.
S. Santamarina-Fojo, A. T. Remaley, E. B. Neufeld, and H.B. Brewer Jr., “Regulation and intracellular trafficking of the ABCA1 transporter,” Journal of Lipid Research, vol. 42, no. 9, pp. 1339–1345, 2001.
J. M. Timmins, J. Lee, E. Boudyguina et al., “Targeted inactivation of hepatic Abca1 causes profound hypoalphalipoproteinemia and kidney hypercatabolism of apoA-I,” Journal of Clinical Investigation, vol. 115, no. 5, pp. 1333–1342, 2005.
L. R. Brunham, R. R. Singaraja, M. Duong, et al., “Tissue-specific roles of ABCA1 influence susceptibility to atherosclerosis,” Arteriosclerosis, Thrombosis, and Vascular Biology, vol. 29, no. 4, pp. 548–554, 2009.
L. R. Brunham, J. K. Kruit, T. D. Pape et al., “β-cell ABCA1 influences insulin secretion, glucose homeostasis and response to thiazolidinedione treatment,” Nature Medicine, vol. 13, no. 3, pp. 340–347, 2007.
J. Lee, A. Shirk, J. F. Oram, S. P. Lee, and R. Kuver, “Polarized cholesterol and phospholipid efflux in cultured gall-bladder epithelial cells: evidence for an ABCA1-mediated pathway,” Biochemical Journal, vol. 364, no. 2, pp. 475–484, 2002.
J. McNeish, R. J. Aiello, D. Guyot et al., “High density lipoprotein deficiency and foam cell accumulation in mice with targeted disruption of ATp-binding cassette transporter-1,” Proceedings of the National Academy of Sciences of the United States of America, vol. 97, no. 8, pp. 4245–4250, 2000.
M. A. Wollmer, J. R. Streffer, D. Lütjohann et al., “ABCA1 modulates CSF cholesterol levels and influences the age at onset of Alzheimer？s disease,” Neurobiology of Aging, vol. 24, no. 3, pp. 421–426, 2003.
P. D. Sundar, E. Feingold, R. L. Minster, S. T. DeKosky, and M. I. Kamboh, “Gender-specific association of ATP-binding cassette transporter 1 (ABCA1) polymorphisms with the risk of late-onset Alzheimer's disease,” Neurobiology of Aging, vol. 28, no. 6, pp. 856–862, 2007.
M. Ota, T. Fujii, K. Nemoto et al., “A polymorphism of the ABCA1 gene confers susceptibility to schizophrenia and related brain changes,” Progress in Neuro-Psychopharmacology and Biological Psychiatry, vol. 35, no. 8, pp. 1877–1883, 2011.
M. F. Rosenberg, R. Callaghan, R. C. Ford, and C. F. Higgins, “Structure of the multidrug resistance p-glycoprotein to 2.5？nm resolution determined by electron microscopy and image analysis,” The Journal of Biological Chemistry, vol. 272, no. 16, pp. 10685–10694, 1997.
M. L. Fitzgerald, A. J. Mendez, K. J. Moore, L. P. Andersson, H. A. Panjeton, and M. W. Freeman, “ATP-binding cassette transporter A1 Contains an NH2-terminal signal anchor sequence that translocates the protein's first hydrophilic domain to the exoplasmic space,” Journal of Biological Chemistry, vol. 276, no. 18, pp. 15137–15145, 2001.
F. Scheffel, U. Demmer, E. Warkentin, A. Hülsmann, E. Schneider, and U. Ermler, “Structure of the ATPase subunit CysA of the putative sulfate ATP-binding cassette (ABC) transporter from Alicyclobacillus acidocaldarius,” FEBS Letters, vol. 579, no. 13, pp. 2953–2958, 2005.
R. S. Kiss, N. Kavaslar, K. Okuhira, et al., “Genetic etiology of isolated low HDL syndrome: incidence and heterogeneity of efflux defects,” Arteriosclerosis, Thrombosis, and Vascular Biology, vol. 27, no. 5, pp. 1139–1145, 2007.
K. Alrasadi, I. L. Ruel, M. Marcil, and J. Genest, “Functional mutations of the ABCA1 gene in subjects of French-Canadian descent with HDL deficiency,” Atherosclerosis, vol. 188, no. 2, pp. 281–291, 2006.
M. Mantaring, J. Rhyne, S. Ho Hong, and M. Miller, “Genotypic variation in ATP-binding cassette transporter-1 (ABCA1) as contributors to the high and low high-density lipoprotein-cholesterol (HDL-C) phenotype,” Translational Research, vol. 149, no. 4, pp. 205–210, 2007.
T. L. Slatter, G. T. Jones, M. J. A. Williams, A. M. van Rij, and S. P. A. McCormick, “Novel rare mutations and promoter haplotypes in ABCA1 contribute to low-HDL-C levels,” Clinical Genetics, vol. 73, no. 2, pp. 179–184, 2008.
M. T. Chhabria, B. N. Suhagia, and S. B. Pathik, “HDL elevation and lipid lowering therapy: current scenario and future perspectives,” Frontiers in Cardiovascular Drug Discovery, vol. 1, pp. 32–60, 2010.
C. Albrecht, K. Baynes, A. Sardini et al., “Two novel missense mutations in ABCA1 result in altered trafficking and cause severe autosomal recessive HDL deficiency,” Biochimica et Biophysica Acta, vol. 1689, no. 1, pp. 47–57, 2004.
M. Daimon, T. Kido, M. Baba et al., “Association of the ABCA1 gene polymorphisms with type 2 DM in a Japanese population,” Biochemical and Biophysical Research Communications, vol. 329, no. 1, pp. 205–210, 2005.
R. M. Lawn, D. P. Wade, M. R. Garvin et al., “The Tangier disease gene product ABC1 controls the cellular apolipoprotein-mediated lipid removal pathway,” Journal of Clinical Investigation, vol. 104, no. 8, pp. R25–R31, 1999.
A. von Eckardstein, C. Langer, T. Engel et al., “ATP binding cassette transporter ABCA1 modulates the secretion of apolipoprotein E from human monocyte-derived macrophages,” The FASEB Journal, vol. 15, no. 9, pp. 1555–1561, 2001.
M. R. Hayden, S. M. Clee, A. Brooks-Wilson, J. Genest Jr., A. Attie, and J. J. P. Kastelein, “Cholesterol efflux regulatory protein, Tangier disease and familial high density lipoprotein deficiency,” Current Opinion in Lipidology, vol. 11, no. 2, pp. 117–122, 2000.
R. Frikke-Schmidt, B. G. Nordestgaard, G. B. Jensen, and A. Tybj？rg-Hansen, “Genetic variation in ABC transporter A1 contributes to HDL cholesterol in the general population,” The Journal of Clinical Investigation, vol. 114, no. 9, pp. 1343–1353, 2004.
M. Marcil, L. Yu, L. Krimbou et al., “Cellular cholesterol transport and efflux in fibroblasts are abnormal in subjects with familial HDL deficiency,” Arteriosclerosis, Thrombosis, and Vascular Biology, vol. 19, no. 1, pp. 159–169, 1999.
M. E. Brousseau, G. P. Eberhart, J. Dupuis et al., “Cellular cholesterol efflux in heterozygotes for Tangier disease is markedly reduced and correlates with high density lipoprotein cholesterol concentration and particle size,” Journal of Lipid Research, vol. 41, no. 7, pp. 1125–1135, 2000.
S. M. Clee, J. J. P. Kastelein, M. van Dam et al., “Age and residual cholesterol efflux affect HDL cholesterol levels and coronary artery disease in ABCA1 heterozygotes,” The Journal of Clinical Investigation, vol. 106, no. 10, pp. 1263–1270, 2000.
N. F. Fitz, A. A. Cronican, M. Saleem et al., “Abca1 deficiency affects Alzheimer's disease-like phenotype in human ApoE4 but not in ApoE3-targeted replacement mice,” Journal of Neuroscience, vol. 32, no. 38, pp. 13125–13136, 2012.
E. J. Schaefer, L. A. Zech, D. E. Schwartz, and H. B. Brewer Jr., “Coronary heart disease prevalence and other clinical features in familial high-density lipoprotein deficiency (Tangier disease),” Annals of Internal Medicine, vol. 93, no. 2, pp. 261–266, 1980.
P. Cingolani, A. Platts, L. L. Wang et al., “A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff: SNPs in the genome of Drosophila melanogaster strain w 1118; iso-2; iso-3,” Fly, vol. 6, no. 2, pp. 80–92, 2012.
J. C. Cohen, R. S. Kiss, A. Pertsemlidis, Y. L. Marcel, R. McPherson, and H. H. Hobbs, “Multiple rare alleles contribute to low plasma levels of HDL cholesterol,” Science, vol. 305, no. 5685, pp. 869–872, 2004.
R. R. Singaraja, H. Visscher, E. R. James et al., “Specific mutations in ABCA1 have discrete effects on ABCA1 function and lipid phenotypes both in vivo and in vitro,” Circulation Research, vol. 99, no. 4, pp. 389–397, 2006.
M. L. Fitzgerald, A. L. Morris, J. S. Rhee, L. P. Andersson, A. J. Mendez, and M. W. Freeman, “Naturally occurring mutations in the largest extracellular loops of ABCA1 can disrupt its direct interaction with apolipoprotein A-I,” The Journal of Biological Chemistry, vol. 277, no. 36, pp. 33178–33187, 2002.
A. M. Vaughan, C. Tang, and J. F. Oram, “ABCA1 mutants reveal an interdependency between lipid export function, apoA-I binding activity, and Janus kinase 2 activation,” Journal of Lipid Research, vol. 50, no. 2, pp. 285–292, 2009.
R. Frikke-Schmidt, C. F. Sing, B. G. Nordestgaard, and A. Tybj？rg-Hansen, “Gender- and age-specific contributions of additional DNA sequence variation in the 5′ regulatory region of the APOE gene to prediction of measures of lipid metabolism,” Human Genetics, vol. 115, no. 4, pp. 331–345, 2004.
R. Frikke-Schmidt, B. G. Nordestgaard, P. Schnohr, and A. Tybj？rg-Hansen, “Single nucleotide polymorphism in the low-density lipoprotein receptor is associated with a threefold risk of stroke: a case-control and prospective study,” European Heart Journal, vol. 25, no. 11, pp. 943–951, 2004.
S. M. Clee, A. H. Zwinderman, J. C. Engert et al., “Common genetic variation in ABCA1 is associated with altered lipoprotein levels and a modified risk for coronary artery disease,” Circulation, vol. 103, no. 9, pp. 1198–1205, 2001.
M. V. Reddy, I. Iatan, D. Weissglas-Volkov, et al., “Exome sequencing identifies 2 rare variants for low high-density lipoprotein cholesterol in an extended family,” Circulation: Cardiovascular Genetics, vol. 5, no. 5, pp. 538–546, 2012.
C. L. Wellington, Y. Yang, S. Zhou et al., “Truncation mutations in ABCA1 suppress normal upregulation of full-length ABCA1 by 9-cis-retinoic acid and 22-R-hydroxycholesterol,” Journal of Lipid Research, vol. 43, no. 11, pp. 1939–1949, 2002.
S. Wang, K. Gulshan, G. Brubaker, S. L. Hazen, and J. D. Smith, “ABCA1 mediates unfolding of apolipoprotein AI N terminus on the cell surface before lipidation and release of nascent high-density lipoprotein,” Arteriosclerosis, Thrombosis, and Vascular Biology, vol. 33, no. 6, pp. 1197–1205, 2013.
M. Denis, B. Haidar, M. Marcil, M. Bouvier, L. Krimbou, and J. Genest, “Characterization of oligomeric human ATP binding cassette transporter A1: potential implications for determining the structure of nascent high density lipoprotein particles,” The Journal of Biological Chemistry, vol. 279, no. 40, pp. 41529–41536, 2004.
S. Bertolini, L. Pisciotta, M. Seri et al., “A point mutation in ABC1 gene in a patient with severe premature coronary heart disease and mild clinical phenotype of Tangier disease,” Atherosclerosis, vol. 154, no. 3, pp. 599–605, 2001.
W. S. Kim, A. F. Hill, M. L. Fitzgerald, M. W. Freeman, G. Evin, and B. Garner, “Wild type and tangier disease ABCA1 mutants modulate cellular amyloid-β production independent of cholesterol efflux activity,” Journal of Alzheimer's Disease, vol. 27, no. 2, pp. 441–452, 2011.
Y. Nishida, K. Hirano, K. Tsukamoto et al., “Expression and functional analyses of novel mutations of ATP-binding cassette transporter-1 in Japanese patients with high-density lipoprotein deficiency,” Biochemical and Biophysical Research Communications, vol. 290, no. 2, pp. 713–721, 2002.
N. Wang, D. Lan, M. Gerbod-Giannone et al., “ATP-binding cassette transporter A7 (ABCA7) binds apolipoprotein A-I and mediates cellular phospholipid but not cholesterol efflux,” The Journal of Biological Chemistry, vol. 278, no. 44, pp. 42906–42912, 2003.
S. Ho Hong, J. Rhyne, K. Zeller, and M. Miller, “Novel ABCA1 compound variant associated with HDL cholesterol deficiency,” Biochimica et Biophysica Acta—Molecular Basis of Disease, vol. 1587, no. 1, pp. 60–64, 2002.
M. Walter, S. Kerber, C. Fechtrup, U. Seedorf, G. Breithardt, and G. Assmann, “Characterization of atherosclerosis in a patient with familial high-density lipoprotein deficiency,” Atherosclerosis, vol. 110, no. 2, pp. 203–208, 1994.
R. P. Koldamova, I. M. Lefterov, M. Staufenbiel et al., “The liver X receptor ligaun T0901317 decreases amyloid β production in vitro and in a mouse model of Alzheimer's disease,” The Journal of Biological Chemistry, vol. 280, no. 6, pp. 4079–4088, 2005.
T. L. Slatter, M. J. A. Williams, R. Frikke-Schmidt, A. Tybj？rg-Hansen, I. M. Morison, and S. P. A. McCormick, “Promoter haplotype of a new ABCA1 mutant influences expression of familial hypoalphalipoproteinemia,” Atherosclerosis, vol. 187, no. 2, pp. 393–400, 2006.
R. J. Suetani, B. Sorrenson, J. D. A. Tyndall, M. J. A. Williams, and S. P. A. McCormick, “Homology modeling and functional testing of an ABCA1 mutation causing Tangier disease,” Atherosclerosis, vol. 218, no. 2, pp. 404–410, 2011.
T. Fasano, L. Bocchi, L. Pisciotta, S. Bertolini, and S. Calandra, “Denaturing high-performance liquid chromatography in the detection of ABCA1 gene mutations in familial HDL deficiency,” Journal of Lipid Research, vol. 46, no. 4, pp. 817–822, 2005.
M. E. Brousseau, E. J. Schaefer, J. Dupuis, et al., “Novel mutations in the gene encoding ATP-binding cassette 1 in four tangier disease kindreds,” Journal of Lipid Research, vol. 41, no. 3, pp. 433–441, 2000.
W. Huang, K. Moriyama, T. Koga et al., “Novel mutations in ABCA1 gene in Japanese patients with Tangier disease and familial high density lipoprotein deficiency with coronary heart disease,” Biochimica et Biophysica Acta, vol. 1537, no. 1, pp. 71–78, 2001.
F. Quazi and R. S. Molday, “Differential phospholipid substrates and directional transport by ATP binding cassette proteins ABCA1, ABCA7, and ABCA4 and disease-causing mutants,” The Journal of Biological Chemistry, vol. 288, no. 48, pp. 34414–34426, 2013.
V. Kolovou, G. Kolovou, A. Marvaki et al., “ATP-binding cassette transporter A1 gene polymorphisms and serum lipid levels in young Greek nurses,” Lipids in Health and Disease, vol. 10, article 56, 2011.
R. Frikke-Schmidt, B. G. Nordestgaard, G. B. Jensen, R. Steffensen, and A. Tybj？rg-Hansen, “Genetic variation in ABCA1 predicts ischemic heart disease in the general population,” Arteriosclerosis, Thrombosis, and Vascular Biology, vol. 28, no. 1, pp. 180–186, 2008.
A. Kitjaroentham, H. Hananantachai, A. Tungtrongchitr, S. Pooudong, and R. Tungtrongchitr, “R219K polymorphism of ATP binding cassette transporter A1 related with low HDL in overweight/obese Thai males,” Archives of Medical Research, vol. 38, no. 8, pp. 834–838, 2007.
A. Cenarro, M. Artieda, S. Castillo et al., “A common variant in the ABCA1 gene is associated with a lower risk for premature coronary heart disease in familial hypercholesterolaemia,” Journal of Medical Genetics, vol. 40, no. 3, pp. 163–168, 2003.
D. Tregouet, S. Ricard, V. Nicaud et al., “In-depth haplotype analysis of ABCA1 gene polymorphisms in relation to plasma ApoA1 levels and myocardial infarction,” Arteriosclerosis, Thrombosis, and Vascular Biology, vol. 24, no. 4, pp. 775–781, 2004.
T. M. Morgan, H. M. Krumholz, R. P. Lifton, and J. A. Spertus, “Nonvalidation of reported genetic risk factors for acute coronary syndrome in a large-scale replication study,” Journal of the American Medical Association, vol. 297, no. 14, pp. 1551–1561, 2007.
Y. Li, K. Tang, K. Zhou et al., “Quantitative assessment of the effect of ABCA1 R219K polymorphism on the risk of coronary heart disease,” Molecular Biology Reports, vol. 39, no. 2, pp. 1809–1813, 2012.
Z. Xiao, J. Wang, W. Chen, P. Wang, and H. Zeng, “Association studies of several cholesterol-related genes (ABCA1, CETP and LIPC) with serum lipids and risk of Alzheimer's disease,” Lipids in Health and Disease, vol. 11, article 163, 2012.
E. B. Neufeld, J. A. Stonik, S. J. Demosky Jr. et al., “The ABCA1 transporter modulates late endocytic trafficking: insights from the correction of the genetic defect in Tangier disease,” The Journal of Biological Chemistry, vol. 279, no. 15, pp. 15571–15578, 2004.
A. Sandhofer, B. Iglseder, S. Kaser, E. Morè, B. Paulweber, and J. R. Patsch, “The influence of two variants in the adenosine triphosphate-binding cassette transporter 1 gene on plasma lipids and carotid atherosclerosis,” Metabolism, vol. 57, no. 10, pp. 1398–1404, 2008.
U. Hodoglugil, D. W. Williamson, Y. Huang, and R. W. Mahley, “Common polymorphisms of ATP binding cassette transporter A1, including a functional promoter polymorphism, associated with plasma high density lipoprotein cholesterol levels in Turks,” Atherosclerosis, vol. 183, no. 2, pp. 199–212, 2005.
M. K. Jensen, J. K. Pai, K. J. Mukamal, K. Overvad, and E. B. Rimm, “Common genetic variation in the ATP-binding cassette transporter A1, plasma lipids, and risk of coronary heart disease,” Atherosclerosis, vol. 195, no. 1, pp. e172–e180, 2007.
I. Porchay-Baldérelli, F. Péan, N. Emery et al., “Relationships between common polymorphisms of adenosine triphosphate-binding cassette transporter A1 and high-density lipoprotein cholesterol and coronary heart disease in a population with type 2 diabetes mellitus,” Metabolism: Clinical and Experimental, vol. 58, no. 1, pp. 74–79, 2009.
M. E. Brousseau, M. Bodzioch, E. J. Schaefer et al., “Common variants in the gene encoding ATP-binding cassette transporter 1 in men with low HDL cholesterol levels and coronary heart disease,” Atherosclerosis, vol. 154, no. 3, pp. 607–611, 2001.
T. Harada, Y. Imai, T. Nojiri et al., “A common Ile 823 Met variant of ATP-binding cassette transporter A1 gene (ABCA1) alters high density lipoprotein cholesterol level in Japanese population,” Atherosclerosis, vol. 169, no. 1, pp. 105–112, 2003.
E. J. Schaefer, D. W. Anderson, L. A. Zech et al., “Metabolism of high density lipoprotein subfractions and constituents in Tangier disease following the infusion of high density lipoproteins,” Journal of Lipid Research, vol. 22, no. 2, pp. 217–228, 1981.
M. T. Villarreal-Molina, C. A. Aguilar-Salinas, M. Rodríguez-Cruz et al., “The ATP-binding cassette transporter A1 R230C variant affects HDL cholesterol levels and BMI in the Mexican population: association with obesity and obesity-related comorbidities,” Diabetes, vol. 56, no. 7, pp. 1881–1887, 2007.
C. A. Aguilar-Salinas, S. Canizales-Quinteros, R. Rojas-Martínez et al., “The non-synonymous Arg230Cys variant (R230C) of the ATP-binding cassette transporter A1 is associated with low HDL cholesterol concentrations in Mexican adults: a population based nation wide study,” Atherosclerosis, vol. 216, no. 1, pp. 146–150, 2011.
S. H. Hong, J. Rhyne, K. Zeller, and M. Miller, “ABCA1Alabama: a novel variant associated with HDL deficiency and premature coronary artery disease,” Atherosclerosis, vol. 164, no. 2, pp. 245–250, 2002.
S. Bungert, L. L. Molday, and R. S. Molday, “Membrane topology of the ATP binding cassette transporter ABCR and its relationship to ABC1 and related ABCA transporters: identification of N-linked glycosylation sites,” The Journal of Biological Chemistry, vol. 276, no. 26, pp. 23539–23546, 2001.