A functional ABCA1 gene variant is associated with low HDL-cholesterol levels and shows evidence of positive selection in Native Americans

Abstract

It has been suggested that the higher susceptibility of Hispanics to metabolic disease is related to their Native American heritage. A frequent cholesterol transporter ABCA1 (ATP-binding cassette transporter A1) gene variant (R230C, rs9282541) apparently exclusive to Native American individuals was associated with low high-density lipoprotein cholesterol (HDL-C) levels, obesity and type 2 diabetes in Mexican Mestizos. We performed a more extensive analysis of this variant in 4405 Native Americans and 863 individuals from other ethnic groups to investigate genetic evidence of positive selection, to assess its functional effect in vitro and to explore associations with HDL-C levels and other metabolic traits. The C230 allele was found in 29 of 36 Native American groups, but not in European, Asian or African individuals. C230 was observed on a single haplotype, and C230-bearing chromosomes showed longer relative haplotype extension compared with other haplotypes in the Americas. Additionally, single-nucleotide polymorphism data from the Human Genome Diversity Panel Native American populations were enriched in significant integrated haplotype score values in the region upstream of the ABCA1 gene. Cells expressing the C230 allele showed a 27% cholesterol efflux reduction (P< 0.001), confirming this variant has a functional effect in vitro. Moreover, the C230 allele was associated with lower HDL-C levels (P = 1.77 x 10(-11)) and with higher body mass index (P = 0.0001) in the combined analysis of Native American populations. This is the first report of a common functional variant exclusive to Native American and descent populations, which is a major determinant of HDL-C levels and may have contributed to the adaptive evolution of Native American populations.

Figure 1.

Frequency distribution of the C230 allele [(%) black-shaded area] in Native American, European, Asian and African populations. C230 frequency data from populations with an asterisk were obtained from previous reports (8–11 and HapMap and SNP500CANCER databases).

Figure 2.

MP-based network describing the evolutionary relationships of 11 distinct haplotypes. Native American and Mexican-Mestizo haplotypes were established in trios; the remainder were inferred from HapMap groups. Each haplotype is represented by a circle whose area reflects the overall number of copies observed and whose color-coding indicates the frequency of the haplotype in the HapMap groups and Native Americans and Mexican Mestizos. Line length is proportional to the number of differences between haplotypes. Non-filled circles represent non-sampled haplotypes reconstructed by the MP algorithm as evolutionary intermediaries between observed haplotypes. The phylogenetic reconstruction uncovered two major lineages (haplogroups A and B) defined by the non-synonymous polymorphism R219K. The R230C variant occurred on haplogroup B characterized by the ancestral R219 allele, which is frequent in Europe, Asia and America but infrequent in African populations. The C230 allele was found in only one genetic block in all Mexican-Mestizo and Native American individuals analyzed.

Figure 3.

EHH and REHH of ABCA1/R230C and 23 additional SNPs × physical distance in Native American individuals. C230-bearing chromosomes (in gray) appear to have greatly extended LD compared with non-C230-bearing chromosomes (in black). Gray vertical lines in the upper track represent genotyped SNPs, and boxes indicate annotated genes.

Figure 4.

iHS values for individual SNPs flanking the ABCA1 region (2 Mb) × physical distance (top panel) and P-values for the composite likelihood ratio (CLR) test based on a 31-SNP sliding window analysis to detect local regions enriched for high iHS values (bottom panel) in the combined Native American sample. Filled circles indicate P-values <0.05 genome-wide significance level.

Figure 5.

Functional characterization of the ABCA1/R230C variant by lipid efflux assay. (A) Polyclonal stable cell lines expressing the ABCA1 wild-type (WT), variant R230C and mutant M1091T (known defective in lipid efflux) were generated, and efflux activity for cholesterol was performed as described in Materials and Methods. Data represent mean ± SD of two to five experiments as a percent of the ApoA1-dependent efflux induced by wild-type ABCA1. Each assay was performed in triplicate. *P < 0.001. (B) The expression of WT, variant R230C and mutant M1091T ABCA1 protein in Flp-In cells was assessed by western immunoblotting.