An increased level of low-density lipoprotein (LDL) cholesterol, the so-called “bad cholesterol”, in the circulation is a primary risk factor for the development of cardiovascular heart disease. Our lab’s primary research interests are cellular and molecular mechanisms that regulate cholesterol homeostasis in the body and ultimately blood LDL-cholesterol levels.
Clearance of LDL from the circulation requires the LDL receptor, a protein on the cell surface that binds to LDL particles with high affinity and mediates their uptake into cells, mainly in the liver. Since the liver is the primary means for LDL clearance, increased liver LDLR protein expression is a highly desirable goal for therapies aimed at lowering cardiovascular disease risk. Indeed, the efficacy of the widely prescribed statin drugs is ascribed to their ability to increase liver LDL receptors at the gene transcriptional level. However, many patients undergoing statin therapy do not reach therapeutic goals for cholesterol lowering or suffer unacceptable secondary effects, highlighting the need for improved and/or alternate means for increasing liver LDL receptor function.
On this page
Publications
See current publications list at PubMed.
Selected publications:
1. Thomas A. Lagace (2009) PCSK9 and heart disease: quieting an outdated metabolic moderator. Clin. Lipidology 4(4), pp. 407-410.
2. 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.
3. 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.
4. 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.
5. 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.
6. 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.
Staff
Lab Director
Tom Lagace, PhD
Research Assistant
Tanja Francetic, MSc
Graduate Student
Samantha Sarkar, BSc
Focus
The removal of LDL from the circulation occurs mainly in liver via the LDL receptor (LDLR), a cell surface glycoprotein that binds to LDL particles with high affinity and mediates their endocytosis. Internalized LDL particles are degraded in lysosomes and liberated cholesterol is then used for the synthesis of membranes, steroid hormones, lipoproteins and bile acids.
This process is subject to a negative feedback mechanism that maintains tight control of cellular cholesterol levels. Cholesterol influx results in transcriptional suppression of genes encoding cholesterol biosynthetic enzymes and LDLR, thus preventing potentially toxic cholesterol over-accumulation. Conversely, when cholesterol levels are depleted the expression of these same genes is stimulated.
Cholesterol-lowering statin drugs modulate this regulatory circuit by directly inhibiting the rate-controlling enzyme in the cholesterol biosynthetic pathway, resulting in increased LDLR expression, increased LDL uptake by the liver, and lowered plasma LDL levels. Thus, negative feedback control in the liver ultimately dictates plasma LDL-cholesterol levels and associated cardiovascular heart disease risk. We are studying mechanisms that affect cholesterol uptake and trafficking in cells with an emphasis on how these processes affect negative feedback control of cholesterol metabolism.
Projects
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 harbouring 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.
Gallery
Protein Purification and Analysis Core Equipment
Left to right: AKTA Purifier FPLC (GE Healthcare), Profinia Protein Purification (Bio-Rad), Licor Odyssey Infrared Imager (Licor Biosciences)
Available to Heart Institute investigators. Contact Tom Lagace for information.