Lagace, Thomas - University of Ottawa - Department of Biochemistry, Microbiology and Immunology

研究领域

Studies of PCSK9-Mediated LDL Receptor Degradation An aspect of negative feedback control of cholesterol metabolism is the co-regulation of genes encoding the LDLR and PCSK9 (proprotein convertase subtilisin/kexin type-9), a secreted protease that promotes LDLR degradation in liver. Importantly, loss-of-function mutations in PCSK9 have been identified in the human population and are associated with lowered plasma LDL levels and greatly decreased incidence of cardiovascular disease. This exciting finding validates PCSK9 as a therapeutic target for LDL lowering. In previous work we have shown that secreted PCSK9 is active in the circulation, and that PCSK9 binds directly to LDLRs on the surface of hepatic cells leading to LDLR degradation in the endosomal/lysosomal compartment. Surprisingly, PCSK9’s protease activity is not required for LDLR degradation - instead it acts as a molecular chaperone to interfere with LDLR recycling. We have recently identified the pertinent regions on both LDLR and PCSK9 involved in direct binding between these proteins. Our future goals are to identify and characterize regulatory mechanisms that affect both the initial PCSK9:LDLR interaction as well as the downstream cellular degradative pathway utilizing cell-based approaches as well as protein-protein interaction studies with purified protein components. Mechanisms of intracellular cholesterol trafficking The protein machinery that ultimately regulates negative feedback control of cholesterol metabolism is located in the endoplasmic reticulum (ER) and is regulated by cholesterol content in this organelle. Cholesterol trafficking pathways that deliver LDL-derived free cholesterol from lysosomes to the ER play a critical role in this process, yet these pathways remain poorly understood. Phosphatidylcholine (PC), the most abundant phospholipid in cell membranes, can positively influence the incorporation and bilateral movement of cholesterol in membrane bilayers. Within cells, PC and cholesterol content in membranes are maintained within narrow ratios. Using Chinese hamster ovary (CHO) cell lines harboring altered cholesterol and/or PC metabolic genes we are studying how altered cholesterol/PC ratios in the cell influence the trafficking of LDL-derived free cholesterol from the lysosomal compartment to other membrane sites, including the plasma membrane and the ER.

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Thomas A. Lagace (2009) PCSK9 and heart disease: quieting an outdated metabolic moderator. Clin. Lipidology 4(4), pp. 407-410. Markey C. McNutt, Hyock Joo Kwon, Chiyuan Chen, Justin R. Chen, Jay D. Horton and Thomas A. Lagace (2009) Antagonism of Secreted PCSK9 Increases Low-Density Lipoprotein Receptor Expression in HepG2 Cells. J. Biol. Chem. 284, pp. 10561-10570. Hyock Joo Kwon, Thomas A. Lagace, Markey C. McNutt, Jay D. Horton, and Johann Deisenhofer (2008) Molecular basis for LDL receptor recognition by PCSK9. Proc. Nat. Acad. Sci. 105, pp.1820-1825. Markey C. McNutt, Thomas A. Lagace, and Jay D. Horton (2007) Catalytic Activity is Not Required for Secreted PCSK9 to Reduce LDL Receptors in HepG2 Cells. J. Biol. Chem. 282, pp. 20799-20803. Thomas A. Lagace, David E. Curtis, Rita Garuti, Markey C. McNutt, Sahng Wook Park, Heidi B. Prather, Norma N. Anderson, Y. K. Ho, Robert E. Hammer, and Jay D. Horton (2006) Secreted PCSK9 Decreases LDL Receptors in Hepatocytes and in Livers of Parabiotic Mice. J. Clin. Invest. 116(11) pp. 2995-3005. Thomas A. Lagace and Neale D. Ridgway (2005) The Rate-limiting Enzyme in Phosphatidylcholine Synthesis Regulates Proliferation of the Nucleoplasmic Reticulum. Mol. Biol. Cell 16(3) pp.1120-1130.