Regulation of Hepatic Gluconeogenesis by the CREB:TORC2 Pathway

描述（由申请方提供）：空腹期间，循环胰高血糖素升高通过CREB辅激活因子TORC 2诱导肝细胞生成程序促进肝葡萄糖输出。在进食过程中，TORC 2被隔离在细胞质中，响应于禁食信号而易位到细胞核，在那里它与Forkhead转录因子FOXO 1一起触发促凋亡基因表达。初步研究表明，TORC 2在早期禁食期间通过P300依赖性乙酰化被瞬时激活，此时它通过与组蛋白甲基转移酶复合物的结合刺激促凋亡程序。TORC 2沉默通过SIRT 1介导的脱乙酰化过程中长时间禁食，当FOXO 1被激活。提出了三个目的：在目的I中，将确定P300在早期禁食期间通过乙酰化增强TORC 2活性中的作用。将鉴定TORC 2中的乙酰化位点，并将评价P300在催化TORC 2乙酰化中的重要性。还将探索NAD+依赖性脱乙酰酶SIRT 1在长时间禁食期间通过脱乙酰化沉默TORC 2中的作用。在目的II中，将评估Ser/Thr激酶SIK 2在通过P300磷酸化调节肝TORC 2活性中的重要性。将鉴定P300中的SIK 2磷酸化位点，并分析其在破坏P300：TORC 2相互作用从而降低TORC 2乙酰化中的作用。在目标III中，将确定TORC 2相关组蛋白甲基转移酶（HMT）复合物在禁食期间介导诱导促凋亡程序中的作用。TORC 2对于HMT复合物的募集和禁食期间组蛋白甲基化超过致突变启动子的重要性将通过TORC 2或HMT组分的耗尽以及HMT相互作用缺陷的突变TORC 2蛋白的表达来确定。还将通过鉴定和突变TORC 2中的相关位点来评估HMT通过甲基化TORC 2在调节致肿瘤基因表达中的潜在作用。综上所述，拟议的研究将提供对通过葡萄糖平衡所需的共激活剂调节空腹代谢的调节途径的深入了解，该共激活剂有助于糖尿病高血糖症。这些结果可能会导致识别新的分子靶点，用于开发改善胰岛素抵抗个体血糖控制的治疗性化合物。公共卫生相关性：胰腺激素胰高血糖素通过打开一个称为TORC 2的基因开关来维持禁食期间的循环葡萄糖水平，该基因开关增加肝脏中的葡萄糖产生;胰岛素通过关闭TORC 2开关来防止进食期间血糖异常升高。胰高血糖素和胰岛素通过一组酶对TORC 2开关产生这些相反的作用，这些酶引起TORC 2蛋白质的不同化学变化。通过表征TORC 2中的这些化学变化，并了解它们如何改变这种开关触发葡萄糖产生的能力，我们的研究可能会导致治疗糖尿病患者的新疗法。

DESCRIPTION (provided by applicant): During fasting, elevations in circulating pancreatic glucagon promote hepatic glucose output through induction of the gluconeogenic program by the CREB coactivator TORC2. Sequestered in the cytoplasm during feeding, TORC2 translocates to the nucleus in response to fasting signals, where it triggers gluconeogenic gene expression in concert with the Forkhead transcription factor FOXO1. Preliminary studies indicate that TORC2 is transiently activated through P300-dependent acetylation during early fasting, when it stimulates the gluconeogenic program via an association with a histone methyl-transferase complex. TORC2 is silenced through SIRT1-mediated deacetylation during prolonged fasting, when FOXO1 is reciprocally activated. Three Aims are proposed: In Aim I, the role of P300 in augmenting TORC2 activity through acetylation during early fasting will be determined. Acetylation sites in TORC2 will be identified, and the importance of P300 in catalyzing TORC2 acetylation will be evaluated. The role of the NAD+ dependent deacetylase SIRT1 in silencing TORC2 through deacetylation during prolonged fasting will also be explored. In Aim II, the importance of the Ser/Thr kinase SIK2 in modulating hepatic TORC2 activity through phosphorylation of P300 will be evaluated. SIK2 phosphorylation sites in P300 will be identified, and their role in disrupting the P300:TORC2 interaction and thereby reducing TORC2 acetylation will be analyzed. In Aim III, the role of a TORC2 associated histone methyl-transferase (HMT) complex in mediating induction of the gluconeogenic program during fasting will be determined. The importance of TORC2 for recruitment of HMT complexes and for histone methylation over gluconeogenic promoters during fasting will be determined, by depletion of TORC2 or HMT components, and by expression of HMT interaction-defective mutant TORC2 proteins. The potential role of HMTs in modulating gluconeogenic gene expression by methylating TORC2 will also be evaluated through identification and mutation of relevant sites in TORC2. Taken together, the proposed studies will provide insight into regulatory pathways that modulate fasting metabolism through a coactivator that is required for glucose balance and that contributes to hyperglycemia in diabetes. The results may lead to the identification of new molecular targets for the development of therapeutic compounds that improve glucose control in insulin resistant individuals. PUBLIC HEALTH RELEVANCE: The pancreatic hormone glucagon maintains circulating glucose levels during fasting by turning on a genetic switch, called TORC2, that increases glucose production in the liver; insulin protects against abnormal elevations in blood glucose during feeding by turning off the TORC2 switch. Glucagon and insulin exert these opposing effects on the TORC2 switch through a group of enzymes that cause different chemical changes in the TORC2 protein. By characterizing these chemical changes in TORC2 and understanding how they modify the ability for this switch to trigger glucose production, our studies may lead to new therapies for the treatment of diabetic patients.