Rare variant in scavenger receptor BI raises HDL cholesterol and increases risk of coronary heart disease

Abstract

Scavenger receptor BI (SR-BI) is the major receptor for high-density lipoprotein (HDL) cholesterol (HDL-C). In humans, high amounts of HDL-C in plasma are associated with a lower risk of coronary heart disease (CHD). Mice that have depleted Scarb1 (SR-BI knockout mice) have markedly elevated HDL-C levels but, paradoxically, increased atherosclerosis. The impact of SR-BI on HDL metabolism and CHD risk in humans remains unclear. Through targeted sequencing of coding regions of lipid-modifying genes in 328 individuals with extremely high plasma HDL-C levels, we identified a homozygote for a loss-of-function variant, in which leucine replaces proline 376 (P376L), in SCARB1, the gene encoding SR-BI. The P376L variant impairs posttranslational processing of SR-BI and abrogates selective HDL cholesterol uptake in transfected cells, in hepatocyte-like cells derived from induced pluripotent stem cells from the homozygous subject, and in mice. Large population-based studies revealed that subjects who are heterozygous carriers of the P376L variant have significantly increased levels of plasma HDL-C. P376L carriers have a profound HDL-related phenotype and an increased risk of CHD (odds ratio = 1.79, which is statistically significant).

Fig. 1. HDL composition and functionality in a SCARB1 P376L homozygote, heterozygous carriers, and controls

(A) FPLC fractionation of plasma lipoproteins from the P376L homozygote subject (red) and from a control with normal HDL-C. (B) Cholesterol, apoA-I, and apoA-II content in total HDL. (C) Free cholesterol (FC) and esterified cholesterol (CE) in total HDL (left) and the FC/CE ratio in total HDL (right). (D) HDL subclass concentrations after separation by density-gradient ultracentrifugation. (E) ApoA-I content in the same HDL subclasses. (F) ApoC-III content in the same HDL subclasses. (G) Cholesterol efflux capacity from macrophages of the THP-1 cell line. All data are reported as means ± SD.

Fig. 2. SCARB1 P376L is a null variant in vitro and in vivo

(A) [³H] Cholesterol ether (CEt) uptake (left) and selective cholesterol uptake from HDL (right) in iPSC-derived HLCs from the P376L homozygote versus a noncarrier control. Cells were incubated with [³H]CEt and ¹²⁵I-labeled tyramine cellobiose (TC) dual-labeled human HDL. All values are normalized to relative ALB gene expression in each cell line. All data represent mean values for wells of respective cell lines ± SD. (B) Plasma HDL cholesterol levels before and 12 days after AAV administration to Scarb1 KO mice. (C) [³H]Cholesterol ether (CEt) clearance (left) and fractional catabolic rate (right) from plasma of Scarb1 KO mice injected with null or SR-BI AAVs after administration of [³H]CE/¹²⁵I-labeled TC dual-labeled human HDL. (D) ¹²⁵I-labeled TC clearance (left) and fractional catabolic rate (right) from plasma after administration of dual-labeled HDL. (E) Selective cholesterol uptake in mice expressing null, SR-BI WT, or P376L measured by relative differences in ³H- and ¹²⁵I-labeled fractional catabolic rates. (F) Sensitivity to Endo-H in P376L homozygous versus noncarrier iPSC-derived HLCs. Cell lysates of each genotype were treated with Endo-H to remove complex N-linked glycans from mature forms of proteins and then immunoblotted for SR-BI. Molecular weights of different forms of SR-BI after Endo-H treatment are given on the left. (G) SR-BI Endo-H sensitivity from liver lysates from mice expressing null, SR-BI WT, or SR-BI P376L AAV. Lysates were treated with Endo-H, followed by immunoblotting for SR-BI. Molecular weights of different forms of SR-BI after Endo-H treatment are given on the left. (A) Mean values for wells of respective cell lines ± SD; [(B) to (E)] means ± SD for each of the three groups. *P < 0.05; ** P < 0.01; ***P <0.001 by analysis of variance (ANOVA) [(B) and (C)]; plasma clearance, unpaired t test (E).