Abstract 830: The Proprotein Convertase PCSK9 Induces The Degradation Of LDLR And Its Closest Family Members VLDLR And ApoER2
Introduction: The proprotein convertase PCSK9 gene is the third locus implicated in familial hypercholesteromia, emphasizing its role in cardiovascular diseases. Loss of function mutations and gene disruption of PCSK9 resulted in a higher clearance of plasma low density lipoprotein cholesterol (LDL-C), likely due to a lower degradation of the liver LDL receptor (LDLR).
Hypothesis: PCSK9 enhances the degradation of LDLR and its closest family members ApoER2 and VLDLR.
Methods:  Co-expression of PCSK9 and its membrane-bound chimeras together with LDLR, VLDLR or ApoER2 was analyzed by Western blots in various cells lines. Additionally, 3T3 cells stably expressing either VLDLR or ApoER2 were incubated with recombinant wild type human PCSK9 or its gain of function mutant D374Y.  In situ hybridization was used to compare the mRNA expression of PCSK9 to that of VLDLR or ApoER2 in whole body of 1 day old mice.  Protein levels of VLDLR in skeletal muscle of adult mice (wild type, complete PCSK9 knockout, and transgenic V5-tagged PCSK9 overexpressed in hepatocytes) were measured by immunocytochemistry and Western blots.
Results: The data revealed that wild type PCSK9 and more so its natural gain of function mutant D374Y can efficiently degrade VLDLR and ApoER2 either following co-expression or re-internalization of secreted human PCSK9 as described for the LDLR. We next designed membrane-bound PCSK9-chimeras, which demonstrated that as compared to the secreted wild type enzyme, the enhanced intracellular targeting of PCSK9 to late endosomes/lysosomes resulted in a much more efficient degradation of the three receptors. In vivo validation of the enlarged functional activity profile of PCSK9 was achieved by demonstrating that PCSK9 modulates VLDLR levels in skeletal muscles in both Pcsk9 knockout mice and in liver-specific transgenic mice.
Conclusions: These data provided evidence for the endocrine-like function of secreted hepatic PCSK9, which enhances the degradation of the VLDLR in skeletal muscle. Thus, the effect of PCSK9 on these three receptors suggests that this convertase plays a major role in cholesterol and lipid homeostasis, and through the enhanced degradation of the VLDLR it could also reduce the development of adiposity.