E1A Oncoprotein Induced Deregulation of Replication Origin Firing

描述（由申请人提供）：本申请的重点是确定腺病毒（Ad）转化E1 A蛋白以p300结合依赖性方式诱导静止细胞向S期进展的机制。E1 A与细胞激活的ras或病毒编码的E1 B19 K或55 K蛋白协同作用，并通过与p300/CBP和Rb家族蛋白结合，可转化啮齿动物细胞。E1 A与p300/CBP和Rb蛋白在静止细胞中的相互作用导致S期的快速诱导。虽然众所周知Rb蛋白的失活诱导E2 F和S期，但E1 A结合p300的贡献尚未得到彻底研究。以前我们发现p300通过负调节c-Myc阻止静止期细胞过早进入S期。p300与转录因子YY 1和组蛋白去乙酰化酶3在上游YY 1结合位点协同作用，使c-Myc保持在抑制状态，这有助于维持细胞处于G 0期。E1 A通过干扰该机制诱导c-Myc。最近，我们发现，在静止期细胞中，许多与DNA合成和细胞周期进程相关的基因也被p300抑制，并且这些基因以p300结合依赖的方式被E1 A诱导。通过shRNA下调p300或表达野生型E1 A而不是p300结合缺陷型E1 A突变体导致诱导c-Myc，其与S期的快速诱导、DNA损伤反应的激活和S期停滞有关。这些细胞含有增加的细胞DNA复制起点活性。我们现在提出的研究，以确定如何起源的活动是由E1 A在p300结合依赖性的方式解除管制。这些研究有望为E1 A介导的异常S期进程及其在病毒和细胞转化中的作用提供新的线索。 公共卫生相关性：各种机制对细胞生长控制的失调是人类癌症发展的关键步骤。腺病毒编码的E1 A是一种历史悠久的模式癌蛋白。了解其对异常S期进展的影响将促进我们对癌基因如何通过破坏细胞蛋白质的生长抑制途径促进肿瘤转化的理解。

DESCRIPTION (provided by applicant): The focus of this application is to determine the mechanisms by which adenovirus (Ad) transforming E1A protein induces progression to S phase in quiescent cells, in a p300 binding dependent manner. In cooperation with cellular activated ras or virus encoded E1B19K or 55K proteins and by binding to p300/CBP and Rb family proteins, E1A can transform rodent cells. Interactions of E1A with p300/CBP and Rb proteins in quiescent cells result in rapid induction of S phase. While the inactivation of Rb proteins is well known to induce E2F and S phase, the contribution of E1A binding to p300 has not been thoroughly studied. Previously we showed that p300 prevents premature entry of quiescent cells into S phase by negatively regulating c-Myc. p300 cooperates with transcription factor YY1 and histone deacetylase 3 at an upstream YY1 binding site to keep c-Myc in a repressed state and that this contributes to maintaining cells in G0 phase. E1A induces c-Myc by interfering with this mechanism. Recently, we have discovered that a number of additional genes related to DNA synthesis and cell cycle progression are also repressed by p300 in quiescent cells and these genes are induced by E1A in a p300 binding dependent manner. Either downregulation of p300 by shRNAs or expression of wild type E1A but not p300 binding defective E1A mutants leads to the induction of c-Myc that is linked to rapid induction of S phase, activation of DNA damage response and S phase arrest. These cells contain increased cellular DNA replication origin activity. We now propose studies to determine how the origin activity is deregulated by E1A in a p300 binding dependent manner. These studies are expected to shed new light on E1A mediated aberrant S phase progression and its role in viral and cellular transformation. PUBLIC HEALTH RELEVANCE: Deregulation of cell growth control by a variety of mechanisms is a key step in the development of human cancer. Adenovirus encoded E1A is a time honored model oncoprotein. Understanding its effects on aberrant S phase progression will advance our understanding of how an oncogene by disrupting the growth inhibitory pathways of a cellular protein contributes to neoplastic transformation.