Our goal is to determine the molecular nature of certain common controlling mechanism(s) which may exist in regulating the activities of various cholesterogenic enzymes by low density lipoprotein-(LDL) bound cholesterol or its specific analog (25-hydroxycholesterol), and to isolate and identify the putative common controlling factor(s) involved. We focus our attention on our recent observation that inactivation of HMG-CoA reductase, an important cholesterogenic enzyme, via increase of enzyme degradation rate by LDL or by 25-hydroxycholesterol, is a cycloheximide-sensitive process, implying that continuous synthesis of a class of mediator protein(s) may be necessary in mediating the effect of the sterol. The turnover rate of the putative mediator protein(s) was shown to be very rapid (with half-life less than 3 h). We wish to explore this observation in detail. Our proposed experimentations are categorized as follows: (i) We will design various experiments to see if the putative mediator protein(s) plays any role in mediating the suppression effects of 25-hydroxycholesterol on other early cholesterogenic enzyme activities (acetoacetyl CoA thiolase and HMG-CoA synthase). This will further define the functional specificity of the mediator protein(s) in intact cells. (ii) Based on functional specificity, we proposed to develop specific assays to measure the mediator protein(s) activity in vitro, and attempt to partially characterize the protein(s) in vitro. (iii) We propose to isolate and characterize Chinese hamster ovary cell mutants defective in mediator protein(s) activity. The outcome of this research should provide useful informatin in understanding regulation of cholesterol biosynthesis at the molecular level.
| Cadigan, K M; Heider, J G; Chang, T Y (1988) Isolation and characterization of Chinese hamster ovary cell mutants deficient in acyl-coenzyme A:cholesterol acyltransferase activity. J Biol Chem 263:274-82 |
| Chang, C C; Chang, T Y (1986) Cycloheximide sensitivity in regulation of acyl coenzyme A:cholesterol acyltransferase activity in Chinese hamster ovary cells. 2. Effect of sterol endogenously synthesized. Biochemistry 25:1700-6 |
| Chang, C C; Doolittle, G M; Chang, T Y (1986) Cycloheximide sensitivity in regulation of acyl coenzyme A:cholesterol acyltransferase activity in Chinese hamster ovary cells. 1. Effect of exogenous sterols. Biochemistry 25:1693-9 |
