REGULATION OF APOPROTEIN E BIOSYNTHESIS IN HEPATOTYCTES & ASSOC W/LIPOPROTEIN

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In this project, the three aspects of apoprotein E availability will be studied: the determinants of apoprotein E association with lipoprotein, the post-transcriptional regulation of apoprotein E secretion as stimulated by exogenous lipoproteins and the transcriptional regulation of apoprotein E production in hepatocytes. Apoprotein E associates with triglyceride- rich lipoproteins (VLDL and chylomicron remnants) and with larger HDL particles. The apoprotein sequences that determine these associations will be studied using domain specific mutagenesis and expression in hepatocytes and in heterogeneous systems (baculovirus). Mutant apoprotein E association with nascent and mature VLDL and HDL will be examined. HDL and LDL stimulates apoprotein E production 4-5 fold in steroidogenic cells, without effecting apoprotein E gene transcription. Mechanisms of this stimulation will be studied in a steroidogenic cell line. The domains of apoprotein E required for this effect will be delineated. Finally, the effects of cholesterol loading of hepatocytes and of modifying intracellular sterol balance on the transcription of the apoprotein E gene will be explored in detail. Two agonists -progesterone and 25 hydroxycholesterol will be studied. Efforts will be made to define the cis-acting sequences of apoprotein E gene and the trans-acting factors responsible for this regulation.