Post-transcriptional Regulation of Low Density Lipoprotein Receptor Protein by Proprotein Convertase Subtilisin/Kexin Type 9a in Mouse Liver

Abstract

Lipid homeostasis is transcriptionally regulated by three DNA-binding proteins, designated sterol regulatory element-binding protein (SREBP)-1a, -1c, and -2. Oligonucleotide arrays hybridized with RNA made from livers of transgenic SREBP-1a, transgenic SREBP-2, and SREBP cleavage-activating protein knockout mice recently identified 33 genes regulated by SREBPs in liver, four of which had no known connection to lipid metabolism. One of the four genes was PCSK9, which encodes proprotein convertase subtilisin/kexin type 9a, a protein that belongs to the proteinase K subfamily of subtilases. Mutations in PCSK9 are associated with an autosomal dominant form of hypercholesterolemia. Here, we demonstrate that hepatic overexpression of either wild-type or mutant PCSK9 in mice results in hypercholesterolemia. The hypercholesterolemia is due to a post-transcriptional event causing a reduction in low density lipoprotein (LDL) receptor protein prior to the internalization and recycling of the receptor. Overexpression of PCSK9 in primary hepatocytes and in mice lacking the LDL receptor does not alter apolipoprotein B secretion. These data are consistent with PCSK9 affecting plasma LDL cholesterol levels by altering LDL receptor protein levels via a post-transcriptional mechanism.Plasma LDL 1 cholesterol concentrations are determined by the relative rates of VLDL and LDL production by the liver and the rate of LDL uptake via hepatic LDL receptors (LDLRs) (1, 2). VLDL secretion from hepatocytes is positively correlated with rates of hepatic lipid synthesis (3). Genes required for cholesterol and triglyceride biosynthesis and, thus, VLDL production are regulated by three sterol regulatory element-binding proteins (SREBPs), SREBP-1a, SREBP-1c, and SREBP-2 (4, 5). SREBPs also are the principal transcriptional regulators of the LDL receptor gene, which clears apoB-containing lipoproteins, such as VLDL and LDL, from the plasma (5).To identify genes regulated by SREBPs, we used oligonucleotide arrays hybridized with RNA from livers of mice that overexpressed SREBPs (transgenic for SREBP-1a or transgenic for SREBP-2) and that lacked all SREBPs as a result of deleting SCAP, an escort protein required for SREBP activation (5). With this physiologic filter, 33 genes were identified that were increased in the transgenic livers and decreased in the SCAPdeficient livers. Four of these 33 genes had no known function. One of these four genes was Pcsk9, which encodes the proprotein convertase subtilisin/kexin type 9a, also designated NARC-1 (neural apoptosis-regulated convertase 1). Seidah et al. (6) showed that PCSK9 belongs to the proteinase K subfamily of subtilases. PCSK9 is synthesized first as a soluble zymogen that undergoes autocatalytic intramolecular processing in the ER to produce a prosegment that remains associated with the secreted enzyme.A link between PCSK9 and cholesterol metabolism was established by Abifadel et al. (7), who showed that two missense mutations in PCSK9 were associated with an autosomal dominant form of hype...

Decreased plasma cholesterol and hypersensitivity to statins in mice lacking Pcsk9

“…Overexpression of WT PCSK9 reduced hepatic LDLRs to a similar extent as expression of PCSK9 mutant forms (7). These studies suggested that PCSK9 might function normally to reduce LDLR expression levels in liver.…”

“…Recent studies suggest that hepatic LDLRs also may be posttranscriptionally regulated by proprotein convertase subtilisin͞ kexin type 9a (PCSK9) (5)(6)(7). PCSK9 belongs to the proteinase K subfamily of subtilases, which are proteinases synthesized as soluble zymogens that subsequently undergo autocatalytic cleavage to active enzymes (8).…”

Mutations and polymorphisms in the proprotein convertase subtilisin kexin 9 (PCSK9) gene in cholesterol metabolism and disease

“…Pcsk9 overexpression in wild-type mice caused a two-fold increase in plasma total cholesterol and a five-fold increase in non-HDL cholesterol whereas overexpression of Pcsk9 had no effect on LDLR mRNA levels [Maxwell and Breslow, 2004]. Similarly to wild type, human mutant PCSK9 (p.S127R and p.F216L) overexpressed in the liver of mice leads to hypercholesterolemia due to a dramatic decrease of hepatic LDL receptor levels through a posttranscriptional mechanism [Park et al, 2004]. Conversely, livers of knockout mice lacking PCSK9 (Pcsk9-/-) display increased LDL receptor (but not mRNA) that leads to a decrease in plasma cholesterol levels of 48% compared to wildtype litter mates [Rashid et al, 2005].…”

A Novel Loss of Function Mutation of PCSK9 Gene in White Subjects With Low-Plasma Low-Density Lipoprotein Cholesterol

“…10 -12 In mice the overexpression of wild-type PCSK9, as well as of mutant PCSK9 associated with hypercholesterolemia, resulted in a marked reduction of hepatic LDLreceptor (LDL-r) protein leading to hypercholesterolemia. [13][14][15] Missense mutations of PCSK9 associated with hypercholesterolemia in humans would increase the capacity of PCSK9 to limit the number of LDL-r, thus representing gain of function mutations. In sharp contrast, mice with targeted inactivation of PCSK9 gene (pcsk9 Ϫ/Ϫ ) have an increased number of LDL-r in the liver, increased removal of plasma LDL, and reduced plasma LDL-C. 16 In 2005 Cohen et al 17 found that 2 inactivating mutations of PCSK9 (Y142X and C679X), present in 2% to 2.6% of blacks of the Dallas Heart Study, were associated with 30% to 40% reduction of plasma LDL-C.…”