Secreted PCSK9 decreases the number of LDL receptors in hepatocytes and inlivers of parabiotic mice

Abstract

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a member of the proteinase K subfamily of subtilases that reduces the number of LDL receptors (LDLRs) in liver through an undefined posttranscriptional mechanism. We show that purified PCSK9 added to the medium of HepG2 cells reduces the number of cellsurface LDLRs in a dose-and time-dependent manner. This activity was approximately 10-fold greater for a gain-of-function mutant, PCSK9(D374Y), that causes hypercholesterolemia. Binding and uptake of PCSK9 were largely dependent on the presence of LDLRs. Coimmunoprecipitation and ligand blotting studies indicated that PCSK9 and LDLR directly associate; both proteins colocalized to late endocytic compartments. Purified PCSK9 had no effect on cell-surface LDLRs in hepatocytes lacking autosomal recessive hypercholesterolemia (ARH), an adaptor protein required for endocytosis of the receptor. Transgenic mice overexpressing human PCSK9 in liver secreted large amounts of the protein into plasma, which increased plasma LDL cholesterol concentrations to levels similar to those of LDLR-knockout mice. To determine whether PCSK9 was active in plasma, transgenic PCSK9 mice were parabiosed with wild-type littermates. After parabiosis, secreted PCSK9 was transferred to the circulation of wild-type mice and reduced the number of hepatic LDLRs to nearly undetectable levels. We conclude that secreted PCSK9 associates with the LDLR and reduces hepatic LDLR protein levels.

Versatility in ligand recognition by LDL receptor family proteins: advances and frontiers

“…Recent studies show that the self-cleaved form of PCSK9 binds directly to the LDLR [28]. Biochemical studies using recombinant proteins have revealed that the first EGF-like repeat of the LDLR (EGF-A) suffices for binding of recombinant, purified PCSK9, and that binding to this repeat is calcium dependent [26].…”

“…Additionally, the affinity of PCSK9 for the receptor is enhanced at endosomal pH [26,29]. Moreover, mutations associated with PCSK9 gain of function result in enhanced LDLR clearance from the cell surface [28,30], and at least one mutant, D374Y, exhibits ~25-fold increased affinity for the receptor at neutral pH [28,29]. However, it is still not clear whether the latent proteolytic activity of the enzyme is required for receptor downmodulation.…”

Plasma PCSK9 in Nephrotic Syndrome and in Peritoneal Dialysis: A Cross-sectional Study

“…On the surface of hepatocytes, PCSK9 binds to the LDLR, forming a complex that is internalized and directs LDLR for intracellular degradation. 11 It should be noted that PCSK9 acts as a chaperone to facilitate intracellular degradation of LDLR, and that this role is independent of its enzymatic activity. 12,13 By promoting LDLR degradation, PCSK9 prevents recycling of LDLR to the cell membrane, leading to a post-translational reduction in LDLR expression.…”

Hepatitis C virus and proprotein convertase subtilisin/kexin type 9: a detrimental interaction to increase viral infectivity and disrupt lipid metabolism

“…Specifically, a concentration‐dependent impact of PCSK9 on liver cell CD81 expression has been documented 16, 17. Also, circulating PCSK9 down‐regulates LDLR expression on the hepatocyte surface, thus decreasing LDL catabolism and increasing plasma LDL‐cholesterol (LDL‐C) levels 18, 19. Although there is evidence that PCSK9 decreases VLDLR expression in adipose tissue 20, whether such a down‐regulation may occur in the liver is not established and the potential impact of PCSK9‐mediated change in VLDLR expression on HCV infection is still unknown.…”