The Role of PARP-1 in Hormone-Regulated Transcription

描述(由申请人提供)： 基因表达的协调调控是生物学中的一个基本过程。在这项研究中，我们探讨了多聚腺苷二磷酸核糖聚合酶-1(PARP-1)及其相关因子在依赖染色质的转录调控中的作用，以响应类固醇激素、雌激素和细胞因子肿瘤坏死因子α的细胞信号。PARP-1是一种丰富的染色质相关酶，催化供体烟酰胺腺嘌呤二核苷酸(NAD+)分子在靶蛋白上聚合聚腺苷二磷酸核糖(PAR)链。PARP-1在多种生理和病理系统中发挥关键作用，包括激素信号、代谢、炎症、分化、脂肪形成、癌症发生、发育和衰老。这些研究的长期目标是更好地理解PARP-1及其相关因素调控信号转录的染色质依赖的分子机制。我们的广泛假设是，PARP-1的基因调控活性取决于：(1)局部染色质环境(例如染色质组成、组蛋白修饰)，(2)PARP-1、转录因子(例如ERα、NF-κB)、组蛋白修饰辅助调节因子和染色质成分之间的物理和功能相互作用，(3)PARP-1酶活性的靶标，以及(4)核内NAD+的可获得性。我们将使用一种综合的方法来测试这一假设，因为它与雌激素和肿瘤坏死因子α依赖的基因调控有关，该方法结合了生物化学、分子生物学、细胞生物学、化学生物学和基因组学的一套互补工具。具体地说，我们将确定(1)依赖染色质的PARP-1在目标基因启动子上的定位和活性的分子机制，包括组蛋白变体H2A.Z和H3.3的作用；(2)依赖PARP-1的位点特异性PAR化对染色质调节蛋白的活性和功能的影响，包括组蛋白去甲基酶KDM5B和组蛋白甲基转移酶Ezh2；以及(3)核NAD+合成酶烟酰胺单核苷酸腺基转移酶-1(NMNAT-1)调节PARP-1在目标基因启动子上的酶活性的机制。总之，这些研究将为在染色质背景下PARP-1‘S基因调控活性的分子机制提供新的见解，特别是与雌激素和肿瘤坏死因子α信号转导以及核NAD+生物合成和可获得性有关的分子机制。鉴于PARP-1在人类疾病中的重要作用，以及它潜在的“可药性”，我们的研究可能会带来利用这些因素作为治疗靶点的新方法。

DESCRIPTION (provided by applicant): The coordinated regulation of gene expression is a fundamental process in biology. In this proposal, we explore the role of poly (ADP-ribose) polymerase-1 (PARP-1) and its associated factors in the chromatin- dependent control of transcription in response to cellular signaling by a steroid hormone, estrogen, and a cytokine, tumor necrosis factor alpha (TNFα). PARP-1 is an abundant, chromatin-associated enzyme that catalyzes the polymerization of poly (ADP-ribose) (PAR) chains on target proteins from donor nicotinamide adenine dinucleotide (NAD+) molecules. PARP-1 plays key roles in a variety of physiological and pathological systems, including hormonal signaling, metabolism, inflammation, differentiation, adipogenesis, carcinogenesis, development, and aging. The long-term objective of these studies is to achieve a better understanding of the chromatin-dependent molecular mechanisms underlying signal-regulated transcription by PARP-1 and its associated factors. Our broad hypothesis is that the gene regulatory activities of PARP-1 are determined by: (1) the local chromatin environment (e.g., chromatin composition, histone modifications), (2) physical and functional interactions among PARP-1, transcription factors (e.g., ERα, NF-κB), histone-modifying coregulators, and components of chromatin, (3) the targets of PARP-1 enzymatic activity, and (4) the availability of NAD+ in the nucleus. We will test this hypothesis as it relates to estrogen- and TNFα-dependent gene regulation by using an integrated approach that combines a complementary set of tools from biochemistry, molecular biology, cell biology, chemical biology, and genomics. Specifically, we will determine (1) the chromatin-dependent molecular mechanisms of PARP-1 localization and activity at target gene promoters, including the role of the histone variants H2A.Z and H3.3; (2) the effects of PARP-1-dependent site- specific PARylation on the activity and function of chromatin-regulating proteins, including the histone demethylase KDM5B and the histone methyltransferase Ezh2; and (3) the mechanisms by which NAD+ production by the nuclear NAD+ synthase nicotinamide mononucleotide adenylyltransferase-1 (NMNAT-1) regulates PARP-1 enzymatic activity at target gene promoters. Collectively, these studies will provide new insights into the molecular mechanisms of PARP-1's gene regulatory activities in the context of chromatin, especially in relation to estrogen and TNFα signaling, and nuclear NAD+ biosynthesis and availability. Given the important role of PARP-1 in human disease, as well as its potential "drugability", our studies could lead to new ways to exploit these factors as therapeutic targets.