Regulation of de novo lipogenesis through BAD-dependent glucose signaling

 描述（由申请人提供）：通过BAD依赖性葡萄糖信号传导调节从头脂肪生成进食和禁食状态下肝脏利用、储存和葡萄糖和脂肪产生之间的稳态平衡由激素和营养提示精细控制。在进食状态下，肝脏首先将多余的葡萄糖储存为糖原，然后通过从头脂肪生成储存为脂肪。肝脏如何感知葡萄糖以确定其利用和储存在分子水平上尚未完全了解。新出现的证据表明，葡萄糖衍生的代谢物，包括葡萄糖激酶（GK）的活性，催化肝糖酵解的第一步，通过提供脂质合成的代谢前体以及触发脂肪生成基因的表达来影响从头脂肪生成。从头脂肪生成的葡萄糖调节至少部分依赖于碳水化合物反应元件结合蛋白（ChREBP），这是一种由某些葡萄糖衍生代谢物激活的转录因子。我提出的研究测试BCL-2家族蛋白BAD作为肝葡萄糖信号传导和通过ChREBP从头脂肪生成的上游调节剂的作用。BAD对葡萄糖代谢的调节是通过Ser 155的磷酸化介导的，其导致GK的直接激活。这种相互作用的功能相关性是显而易见的，从观察到BAD缺乏或干扰其磷酸化与肝脏GK活性和糖酵解降低有关。我发现葡萄糖诱导的ChREBP活性和脂肪生成基因表达在Bad -/-肝细胞中减少。这些数据，连同已知的BAD激活GK的能力和GK在ChREBP激活中的相关性，产生了BAD通过其调节GK活性的能力调节肝ChREBP活性和从头脂肪生成的假设。我将通过以下具体目标来检验这一假设：目标1将询问原代肝细胞中BAD修饰的急性和细胞自主效应以及葡萄糖刺激ChREBP转录活性时GK的伴随变化。目的2将确定功能和代谢相关的脂肪生成基因表达的BAD依赖性变化的生物化学测量的从头脂肪生成和葡萄糖依赖性的变化，在原代肝细胞的脂质谱后，遗传或药理学修饰的BAD。目的3将通过检查ChREBP响应葡萄糖的定义的翻译后修饰的改变，包括已知调节ChREBP转录活性的磷酸化和乙酰化，来剖析BAD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BAD和GK依赖性葡萄糖代谢与ChREBP活性和脂肪生成程序之间联系的途径的综合图。了解这一途径将有助于对肝脏葡萄糖传感，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.