The current application is aimed towards the career development of Russell DeBose-Boyd, Ph.D., Instructor of Molecular Genetics at UT Southwestern Medical Center in Dallas, TX. To prepare the candidate for a research career as an independent investigator, a career development plan is described in which a mentored program is undertaken, encompassing an advanced research experience focusing on a specific research project under the guidance of an appropriate mentor. During the later phases of the program, progressive development of the candidate into an independent investigator will be provided. Experiences gained from the study of lipid metabolism in animals will be applied to parasitic organisms in hopes of creating better treatments for infected patients in economically deprived communities of the United States and the Third World. Excess sterols accelerate degradation of HMG-CoA reductase (HMGCR), a major focal point in the regulation isoprenoid biosynthesis in animal cells. The proposed studies are designed to test the hypothesis that interactions between HMGCR and sterols induce conformational changes in the protein, resulting in enhanced degradation of the enzyme. To gain further insight into this process, three specific aims will be pursued:
(Aim 1) in search of residues important for mediating sterol-regulation, the hydrophobic NH2-terminal domain of HMGCR will be mutagenized in a PCR-based strategy. A selection scheme will be developed to identify mutations that render HMGCR refractory to sterol regulation.
(Aim 2) An assay will be developed to detect structural changes in HMGCR upon sterol treatment, employing HMGCR-specific antibodies and a panel of proteases to probe for sterol-induced conformational changes.
(Aim 3) Parallels between sterol regulation of HMGCR and other sterol-responsive proteins will be explored. Although an important component of cellular membranes, over accumulation of cholesterol is toxic because of its insolubility. Excess cholesterol in the bloodstream can deposit in arteries, initiating atherosclerosis, which can lead to heart attacks. A thorough understanding of molecular mechanisms controlling HMGCR levels may provide insights into novel therapies to treat coronary heart disease.
|Lee, Peter C W; Nguyen, Andrew D; Debose-Boyd, Russell A (2007) Mutations within the membrane domain of HMG-CoA reductase confer resistance to sterol-accelerated degradation. J Lipid Res 48:318-27|
|Nguyen, Andrew D; McDonald, Jeffrey G; Bruick, Richard K et al. (2007) Hypoxia stimulates degradation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase through accumulation of lanosterol and hypoxia-inducible factor-mediated induction of insigs. J Biol Chem 282:27436-46|
|Lee, Peter C W; Liu, Pingsheng; Li, Wei-Ping et al. (2007) Amplification of the gene for SCAP, coupled with Insig-1 deficiency, confers sterol resistance in mutant Chinese hamster ovary cells. J Lipid Res 48:1944-54|
|Song, Bao-Liang; DeBose-Boyd, Russell A (2006) Insig-dependent ubiquitination and degradation of 3-hydroxy-3-methylglutaryl coenzyme a reductase stimulated by delta- and gamma-tocotrienols. J Biol Chem 281:25054-61|
|Goldstein, Joseph L; DeBose-Boyd, Russell A; Brown, Michael S (2006) Protein sensors for membrane sterols. Cell 124:35-46|
|Song, Bao-Liang; Sever, Navdar; DeBose-Boyd, Russell A (2005) Gp78, a membrane-anchored ubiquitin ligase, associates with Insig-1 and couples sterol-regulated ubiquitination to degradation of HMG CoA reductase. Mol Cell 19:829-40|
|Lee, Peter C W; Sever, Navdar; Debose-Boyd, Russell A (2005) Isolation of sterol-resistant Chinese hamster ovary cells with genetic deficiencies in both Insig-1 and Insig-2. J Biol Chem 280:25242-9|
|Song, Bao-Liang; Javitt, Norman B; DeBose-Boyd, Russell A (2005) Insig-mediated degradation of HMG CoA reductase stimulated by lanosterol, an intermediate in the synthesis of cholesterol. Cell Metab 1:179-89|
|Sever, Navdar; Lee, Peter C W; Song, Bao-Liang et al. (2004) Isolation of mutant cells lacking Insig-1 through selection with SR-12813, an agent that stimulates degradation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase. J Biol Chem 279:43136-47|
|Song, Bao-Liang; DeBose-Boyd, Russell A (2004) Ubiquitination of 3-hydroxy-3-methylglutaryl-CoA reductase in permeabilized cells mediated by cytosolic E1 and a putative membrane-bound ubiquitin ligase. J Biol Chem 279:28798-806|
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