Antagonism of secreted PCSK9 increases low density lipoprotein receptor expression in HepG2 cells

Markey C. McNutt, Hyock Joo Kwon, Chiyuan Chen, Justin R. Chen, Jay D. Horton, Thomas A. Lagace

Research output: Contribution to journalArticle

106 Citations (Scopus)

Abstract

PCSK9 is a secreted protein that degrades low density lipoprotein receptors (LDLRs) in liver by binding to the epidermal growth factor-like repeat A (EGF-A) domain of the LDLR. It is not known whether PCSK9 causes degradation of LDLRs within the secretory pathway or following secretion and reuptake via endocytosis. Here we show that a mutation in the LDLR EGF-A domain associated with familial hypercholesterolemia, H306Y, results in increased sensitivity to exogenous PCSK9-mediated cellular degradation because of enhanced PCSK9 binding affinity. The crystal structure of thePCSK9-EGF-A(H306Y)complex shows that Tyr-306 forms a hydrogen bond with Asp-374 in PCSK9 at neutral pH, which strengthens the interaction with PCSK9. To block secreted PCSK9 activity, LDLR (H306Y) subfragments were added to the medium of HepG2 cells stably overexpressing wild-type PCSK9 or gain-of-function PCSK9 mutants associated with hypercholesterolemia (D374Y or S127R). These subfragments blocked secreted PCSK9 binding to cell surface LDLRs and resulted in the recovery of LDLR levels to those of control cells. We conclude that PCSK9 acts primarily as a secreted factor to cause LDLR degradation. These studies support the concept that pharmacological inhibition of the PCSK9-LDLR interaction extracellularly will increase hepatic LDLR expression and lower plasma low density lipoprotein levels.

Original languageEnglish (US)
Pages (from-to)10561-10570
Number of pages10
JournalJournal of Biological Chemistry
Volume284
Issue number16
DOIs
StatePublished - Apr 17 2009

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LDL Receptors
Hep G2 Cells
Epidermal Growth Factor
Degradation
Hyperlipoproteinemia Type II
Secretory Pathway
Liver
Endocytosis
Hypercholesterolemia
LDL Lipoproteins
Hydrogen
Hydrogen bonds
Crystal structure
Plasmas
Recovery
Mutation

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Antagonism of secreted PCSK9 increases low density lipoprotein receptor expression in HepG2 cells. / McNutt, Markey C.; Kwon, Hyock Joo; Chen, Chiyuan; Chen, Justin R.; Horton, Jay D.; Lagace, Thomas A.

In: Journal of Biological Chemistry, Vol. 284, No. 16, 17.04.2009, p. 10561-10570.

Research output: Contribution to journalArticle

McNutt, Markey C. ; Kwon, Hyock Joo ; Chen, Chiyuan ; Chen, Justin R. ; Horton, Jay D. ; Lagace, Thomas A. / Antagonism of secreted PCSK9 increases low density lipoprotein receptor expression in HepG2 cells. In: Journal of Biological Chemistry. 2009 ; Vol. 284, No. 16. pp. 10561-10570.
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