Diurnal regulation of MTP and plasma lipids

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DESCRIPTION (provided by applicant): Our goal is to understand molecular mechanisms involved in the diurnal regulation of plasma lipids. We observed that plasma triglyceride and cholesterol in apoB-lipoproteins show diurnal rhythms. These rhythms are changed when animals are subjected to food entrainment or exposed to constant light. Therefore, both food and light induced mechanisms control variations in plasma lipids. Our aim is to explain both these mechanisms at physiological, biochemical and molecular levels. Aim 1: Light-entrained mechanisms controlling diurnal regulation of plasma lipids and tissue MTP levels: Heterodimers of transcription factors Clock and Bmal1 positively regulate circadian rhythms in the light-entrained suprachiasmatic nucleus of the hypothalamus. These transcription factors are also expressed in peripheral tissues. Our hypothesis is that Clock/Bmal1 are critical for light-induced regulation of plasma lipids and MTP expression. To evaluate this, we will measure plasma triglyceride, cholesterol and apoB-lipoproteins in Clock mutant (Clkmt/mt), Bmal1 (Bmal1-/-) knockout (KO) mice and their wild type (WT) siblings. In addition, protein and mRNA levels of genes involved in lipid metabolism and circadian rhythms will be quantified. To highlight the importance of Clock and Bmal1 in the mobilization of exogenous and endogenous lipids by the intestine and liver, respectively, we will study lipoprotein production by these tissues. Experiments will then focus to understand how these transcription factors regulate MTP gene expression. Elements in MTP promoter critical for circadian regulation and transcription factors that bind to these cis elements will be identified. These studies will elucidate key physiological, biochemical, and molecular mechanisms regulated by Clock and Bmal1 that contribute to diurnal rhythms in plasma lipids. In addition, molecular links in the regulation of MTP by Clock/Bmal1 will be established. Aim 2: Mechanisms involved in food-entrained oscillations in plasma lipids and lipoproteins: We hypothesize that Clock, Bmal1 and MTP play critical roles in food-entrainment of plasma lipids and expression of lipid metabolism and clock genes in the liver and intestine. To test this, Clkmt/mt, Bmal1-/-, as well as intestine and hepatic-specific MTP deficient and control mice will be exposed to different food entrainment schedules. Moreover, the effect of food removal and fasting/re-feeding on plasma lipids and tissue expression of candidate genes will be studied. Experiments will be designed to identify molecular mechanisms involved in food- entrainment of MTP expression. The importance of Clock and Bmal1 in the cyclic MTP expression in cells exposed to 2 h feeding will be ascertained. Elements in MTP promoter that respond to food entrainment and transcription factors that bind to these elements will be identified. These studies will provide novel information about the importance of intestinal and hepatic MTP in the entrainment of peripheral clock genes by food. Critical role of Clock and Bmal1 in food-entrained regulation of plasma lipids/lipoproteins will be established. PUBLIC HEALTH RELEVANCE: We propose to unravel light and food entrained mechanisms that contribute to daily changes in plasma lipid levels. Specifically, the role of Clock and Bmal1 as well as liver and intestine-specific microsomal triglyceride transfer protein will be elucidated.