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）是最常见和高致死率的肝癌形式，然而HCC发病机制的分子机制尚不清楚。有趣的是，在HCC患者中经常观察到代谢调节的破坏，这表明它可能在HCC的发生和/或进展中起着致病作用。代谢调节主要依赖于转录控制，我们的实验室已经发现类固醇受体共激活因子（SRC）家族成员是代谢基因转录的主要调节因子。特别是，SRC-2缺失通过异常调节糖异生和胆汁酸输出而扰乱肝脏代谢功能。我的初步数据表明，与SRC-2+/+对照相比，二乙基亚硝胺（DEN）注射后，SRC-2的缺失增加了肝癌的发生。最近，我们还发现，SRC-2缺乏会显著扰乱中央和外周生物钟，导致已知昼夜节律和昼夜代谢基因的异常循环。此外，SRC- 2可以共同激活主要的昼夜异二聚体转录因子复合体Bmal1：时钟；肝脏SRC-2 ChIP-Seq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。先前的研究表明，包括Bmal1、Per1/2和Cry1/2在内的关键昼夜节律基因的缺失会导致能量调节不正常和自发性肝肿瘤的发生。本研究旨在更好地理解SRC-2作为肿瘤抑制因子的总体作用。我计划通过比较携带SRC-2+/+和SRC-2-/-动物肝组织中关键昼夜节律和昼夜节律控制的代谢和应激反应基因的RNA和蛋白质表达，研究SRC-2消融对Specific Aim 1生物钟的分子影响。此外，我将调查血浆葡萄糖、胆汁酸、甘油三酯和肝脏应激标志物在携带SRC-2 +/+和SRC-2-/-动物中的可能差异。我们计划通过共免疫沉淀和染色质免疫沉淀试验进一步研究SRC-2与昼夜节律转录因子的潜在相互作用。特异性目标2将研究慢性昼夜节律中断对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，对应激标志物进行组织学检查，并测量肝脏应激的血浆标志物。随着HCC发病率的增加，我们有必要了解昼夜节律与癌变之间的分子机制。作为一种公认的能量协同调节因子，SRC-2可能作为昼夜节律时钟的主要调节因子和肝脏肿瘤抑制因子，使肝脏代谢与中心时钟同步。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所必需的肝脏肿瘤抑制因子，并预计SRC-2的缺失会破坏昼夜节律时钟，导致代谢信号异常，导致肝脏应激并最终导致肝细胞癌。

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.