Steroid Receptor Coactivator-2 as a key hepatic tumor suppressor and circadian me

描述(申请人提供)：肝细胞癌(HCC)是最常见和高度致命的肝癌形式，但其发病的分子机制仍不清楚。有趣的是，在肝细胞癌患者中经常观察到代谢调节的紊乱，这表明它可能在肝细胞癌的发生和/或进展中起到致病作用。代谢调控主要依赖于转录调控，本实验室发现类固醇受体辅活化子(SRC)家族成员是代谢基因转录的主要调控因子。特别是，SRC-2的缺失通过对糖异生和胆汁酸输出的异常调节扰乱了肝脏的代谢功能。我的初步数据表明，与SRC-2+/+对照相比，SRC-2作为肝肿瘤抑制因子，与SRC-2+/+对照相比，SRC-2缺失会增加注射二乙基亚硝胺(DEN)后的肝癌发生。最近，我们还发现，SRC-2缺乏显著扰乱了中枢和外周的生物钟，导致已知的昼夜代谢基因的异常循环。此外，SRC-2还可以共同激活主要的昼夜节律异二聚体转录因子复合体BMal1：Clock；对肝脏SRC-2芯片序列数据的初步分析表明，SRC-2和BMal1有许多共同的靶点，包括参与肝脏代谢和应激反应途径的靶点。以前的文献表明，包括BMal1、PER1/2和Cry1/2在内的关键昼夜节律基因的丢失会导致能量调节不当和自发的肝肿瘤发生。这项建议旨在更好地了解SRC-2作为肿瘤抑制因子的主要作用。我计划通过比较SRC-2+/+和SRC-2-/-动物肝脏组织中关键的昼夜节律和昼夜控制的代谢和应激反应基因的RNA和蛋白质表达，来研究SRC-2消融对特定目标1的生物钟的分子影响。此外，我还将研究携带SRC-2+/+和SRC-2-/-动物的血糖、胆汁酸、甘油三酯和肝脏应激标志物的可能差异。我们计划通过免疫共沉淀和染色质免疫沉淀实验进一步研究SRC-2与昼夜节律转录因子的潜在相互作用。具体目标2将研究慢性昼夜节律紊乱对SRC-2+/+和SRC-2-/-动物肝脏应激和肿瘤发生的影响，测量肿瘤发生率，进行应激标志物的组织学研究，并测量肝脏应激的血浆标志物。随着肝细胞癌发病率的增加，了解昼夜节律与肿瘤发生的分子机制势在必行。SRC-2作为一种公认的能量协同调节因子，可能是生物钟的主要调节者，也可能是肝脏肿瘤抑制因子，使肝脏代谢与中枢时钟同步。因此，我假设SRC-2是调节昼夜节律转录因子以维持肝脏代谢同步性所必需的重要的肝脏肿瘤抑制因子，并预计SRC-2的缺失扰乱了昼夜节律，导致代谢信号异常，导致肝脏应激和最终的肝细胞癌。

DESCRIPTION (provided by applicant): Hepatocellular carcinoma (HCC) is the most common and highly lethal form of liver cancer, yet the molecular mechanisms underlying HCC pathogenesis remain poorly understood. Interestingly, disruption of metabolic regulation is frequently observed in HCC patients, suggesting that it may play a causative role in HCC initiation and/or progression. Metabolic regulation principally relies on transcriptional control ad our lab has discovered that Steroid Receptor Coactivator (SRC) family members are major regulators of metabolic gene transcription. In particular, SRC-2 deletion perturbs liver metabolic function through aberrant regulation of gluconeogenesis and bile acid export. My preliminary data suggests that SRC-2 acts as a hepatic tumor suppressor with loss of SRC-2 increasing liver carcinogenesis following diethylnitrosamine (DEN) injection as compared to SRC-2+/+ controls. Recently, we also found that SRC-2 deficiency significantly disrupts the central and peripheral circadian clocks causing abnormal cycling of known circadian and diurnal metabolic genes. Additionally, SRC- 2 can co-activate the primary circadian heterodimeric transcription factor complex Bmal1: clock; and preliminary analysis of liver SRC-2 ChIP-Seq data reveals that SRC-2 and Bmal1 share many targets including those involved in hepatic metabolism and stress response pathways. Previous publications indicate that loss of critical circadian genes including Bmal1, Per1/2, and Cry1/2 results in inapt energy regulation and spontaneous liver tumorigenesis. This proposal aims to better understand the overarching effects of SRC-2 as a tumor suppressor. I plan to investigate the molecular impact of SRC-2 ablation on the circadian clock in Specific Aim 1 by comparing RNA and protein expression of key circadian and circadian controlled metabolic and stress response genes from liver tissue in entrained SRC-2+/+ and SRC-2-/- animals. Additionally, I will investigate possible differences in plasma glucose, bile acid, triglyceride, and hepatic stress markers in entrained SRC- 2+/+ and SRC-2-/- animals. We plan to further investigate the potential interactions of SRC-2 with circadian transcription factor via co-immunoprecipitation and chromatin immunoprecipitation assays. Specific Aim 2 will investigate the effect of chronic circadian disruption on hepatic stress and tumorigenesis in SRC-2+/+ and SRC-2-/- animals measuring tumor incidence, performing histology for stress markers, and measuring plasma markers of hepatic stress. With the increasing prevalence of HCC, it is imperative that we understand the molecular mechanisms linking circadian rhythms with carcinogenesis. As a well-established energy coregulator, SRC-2 may serve as a primary regulator of the circadian clock and a hepatic tumor suppressor synchronizing hepatic metabolism with the central clock. Therefore, I hypothesize that SRC-2 is an essential hepatic tumor suppressor necessary for regulation of circadian transcription factors to maintain metabolic synchrony in the liver and expect that loss of SRC-2 disrupts the circadian clock causing aberrant metabolic signaling, leading to hepatic stress and eventual hepatocellular carcinoma.