Regulation of de novo lipogenesis through BAD-dependent glucose signaling

 描述(由申请人提供)：通过不良依赖的葡萄糖调节新生脂肪生成，在进食和禁食状态下肝脏利用、储存和产生葡萄糖和脂肪之间的动态平衡受到激素和营养信号的精确控制。在饲料状态下，肝脏首先以糖原的形式储存多余的葡萄糖，然后通过新生脂肪生成以脂肪的形式储存。肝脏如何感知葡萄糖以决定其利用和储存，在分子水平上还没有完全了解。越来越多的证据表明，葡萄糖代谢产物，包括催化肝脏糖酵解第一步的葡萄糖激酶(GK)的活性，通过为脂肪合成提供代谢前体以及触发成脂基因的表达来影响从头开始的脂肪生成。葡萄糖对新生脂肪生成的调节至少部分依赖于糖反应元件结合蛋白(ChREBP)，这是一种被某些葡萄糖代谢产物激活的转录因子。我提出的研究测试了bcl2家族蛋白BAD作为肝脏葡萄糖信号的上游调节因子和通过ChREBP进行新生脂肪生成的作用。BAD对糖代谢的调节是通过Ser155的磷酸化而实现的，Ser155直接激活GK。这种相互作用的功能相关性很明显，从观察到严重的缺乏或干扰其磷酸化与肝脏GK活性降低和糖酵解有关。我发现葡萄糖诱导的ChREBP活性和造脂基因在Bad-/-肝细胞中的表达减弱。这些数据结合已知的BAD激活GK的能力和GK在ChREBP激活中的相关性，提出了BAD通过调节GK活性来调节肝脏ChREBP活性和从头脂肪生成的假说。我将通过以下具体目标来验证这一假设：目标1将询问原代肝细胞不良修饰的急性和细胞自主效应，以及伴随而来的GK在葡萄糖刺激ChREBP转录活性方面的变化。目的2通过生化测定BAD基因或药物修饰后原代肝细胞新生脂肪生成和葡萄糖依赖的脂谱变化，确定BAD依赖的造脂基因表达变化的功能和代谢相关性。目的3将通过研究葡萄糖对ChREBP翻译后修饰的改变，包括已知的调节ChREBP转录活性的磷酸化和乙酰化，来剖析不良修饰和ChREBP活性之间的机制联系。随着时间的推移，这些研究将提供一幅连接BAD和GK依赖的葡萄糖代谢与ChREBP活性和造脂程序的完整图景。了解这一途径应该会对肝脏的葡萄糖感知、营养利用和储存产生有用的分子见解。

 DESCRIPTION (provided by applicant): Regulation of de novo lipogenesis through BAD-dependent glucose signaling The homeostatic balance between hepatic utilization, storage, and production of glucose and fat in fed and fasted states is exquisitely controlled by hormonal and nutrient cues. In the fed state, the liver stores excess glucose first as glycogen then as fat through de novo lipogenesis. How the liver senses glucose to determine its utilization and storage is not fully understood at the molecular level. Emerging evidence indicates that glucose-derived metabolites, including the activity of glucokinase (GK), which catalyzes the first step of hepatic glycolysis, influence de novo lipogenesis by both providing metabolic precursors for lipid synthesis as well as triggering the expression of lipogenic genes. Glucose regulation of de novo lipogenesis is dependent, at least in part, on the Carbohydrate Response Element Binding Protein (ChREBP), a transcription factor that is activated by certain glucose-derived metabolites. My proposed studies test the role of the BCL-2 family protein BAD as an upstream regulator of hepatic glucose signaling and de novo lipogenesis through ChREBP. BAD's modulation of glucose metabolism is mediated by phosphorylation of Ser155, which leads to direct activation of GK. The functional relevance of this interaction is evident from the observations that BAD deficiency or interference with its phosphorylation is associated with reduced hepatic GK activity and glycolysis. I have found that glucose induction of ChREBP activity and lipogenic gene expression is diminished in Bad -/- hepatocytes. These data, together with the known capacity of BAD to activate GK and GK's relevance in ChREBP activation, give rise to the hypothesis that BAD modulates hepatic ChREBP activity and de novo lipogenesis through its ability to regulate GK activity. I will test this hypothesis through the following specific aims: Aim 1 will interrogate the acute and cell autonomous effect of BAD modifications in primary hepatocytes and the attendant changes in GK on stimulation of ChREBP transcriptional activity by glucose. Aim 2 will determine the functional and metabolic correlates of BAD-dependent changes in lipogenic gene expression by biochemical measurement of de novo lipogenesis and glucose-dependent changes in lipid profiles in primary hepatocytes following genetic or pharmacologic modification of BAD. Aim 3 will dissect the mechanistic link between BAD modifications and ChREBP activity by examining alterations in defined post translational modifications of ChREBP in response to glucose, including phosphorylation and acetylation known to modulate ChREBP transcriptional activity. In the fullness of time, these studies will provide an integrated picture of the pathway connecting BAD and GK- dependent glucose metabolism with ChREBP activity and the lipogenic program. Understanding this pathway should yield useful molecular insights into hepatic glucose sensing, nutrient utilization and storage.