K-Rta as a Novel SUMO and Epigenetic Regulator

描述(由申请人提供)：基因表达的表观遗传调控在真核生物的许多基本生物学过程中发挥着关键作用。我们专注于确定Kaposi肉瘤疱疹病毒(KSHV)生命周期中涉及染色质蛋白翻译后修饰的表观遗传学机制，KSHV是Kaposi肉瘤(KS)的病原体，KS是一种与艾滋病相关的恶性肿瘤。这种病毒在受感染的细胞中表现出潜伏期，在这种潜伏期中只有几个病毒基因被表达。细胞信号通路可以通过调节近100个病毒基因的表达的时间级联来重新激活潜伏细胞来产生病毒。因此，KSHV是研究影响染色质结构和动力学的表观遗传机制的一个非常有吸引力的模型，染色质结构和动力学是真核细胞中调控转录的关键过程。小泛素样修饰物(SUMO)是一种通过与多种靶蛋白共价连接(即SUMO)来调节包括异染色质形成在内的多种细胞过程的蛋白质。与磷酸化一样，相扑作用也是一种翻译后信号分子，向含有相扑相互作用基序(SIM)的下游靶标传递信号；这些靶标包括影响染色质结构的蛋白质，这些染色质结构通常会抑制转录。我们最近的研究表明，病毒的主要转录反式激活因子K-RTA是一种相扑靶向泛素(Ub)连接酶，它能降解相扑修饰的蛋白质。我们推测，这种相扑靶向Ub连接酶功能对于K-RTA通过破坏潜伏感染细胞中病毒Episome周围的抑制环境来启动KSHV复制的裂解(生产)阶段是重要的。这种抑制环境是由病毒潜伏期相关核抗原(LANA)建立的，LANA是一种相扑结合蛋白，将病毒表型与宿主细胞染色体的异染色区捆绑在一起，从而维持病毒的潜伏状态。这一提议将检验这样一种假设，即相扑修饰的调节在KSHV生命周期中通过调节病毒染色质的结构在该病毒的潜伏时间的建立和重新激活中发挥关键作用。在第一个目标中，我们将研究K-RTA作为依赖相扑的泛素连接酶在KSHV复制中的作用。在第二个目标中，K-RTA以相扑依赖的方式作用并影响病毒染色质的表观遗传特征的潜在机制将被分析。综上所述，这些研究将确定K-RTA以及LANA在KSHV复制中的新角色，并将为控制潜伏期和重新激活的翻译后修饰和染色质重塑提供见解。 公共卫生相关性：对KSHV的这项R01拨款申请的研究将对基因表达调控和表观遗传控制产生新的理解。我们的研究将提供有关病毒重新激活机制的基础知识，并可能产生治疗传染病，特别是由这种致瘤病毒引起的癌症的创新方法。

DESCRIPTION (provided by applicant): Epigenetic regulation of gene expression plays a critical role in many fundamental biological processes in eukaryotes. We have focused on defining epigenetic mechanisms, involving post-translation modification of chromatin proteins, in the life cycle of Kaposi's sarcoma herpesvirus (KSHV), which is an etiological agent of Kaposi's sarcoma (KS), an AIDS-associated malignancy. This virus exhibits a latent phase in infected cells in which only a few viral genes are expressed. Cell signaling pathways can reactivate latent cells to produce virus through a temporal cascade that regulates expression of nearly 100 viral genes. Thus, KSHV is a very attractive model to study epigenetic mechanisms that affect chromatin structure and dynamics, which are key processes regulating transcription in eukaryotic cells. The small ubiquitin-like modifier (SUMO) is a protein that regulates a wide variety of cellular processes, including heterochromatin formation, by covalent attachment (i.e. sumoylation) to a diverse array of target proteins. Sumoylation, like phosphorylation, serves as a post-translational signal molecule to transmit signals to down-stream targets containing a SUMO-interacting motif (SIM); these targets include proteins that impact chromatin structure that generally silence transcription. Our recent studies show that the main viral transcriptional transactivator, K-Rta, is a SUMO-targeting ubiquitin (Ub) ligase, which degrades SUMO- modified proteins. We hypothesize that this SUMO-targeting Ub ligase function is important for K-Rta to initiate the lytic (productive) phase of KSHV replication by disrupting the repressive environment surrounding of the viral episome in latently infected cells. This repressive environment is established by the viral latency- associated nuclear antigen (LANA), which is a SUMO-binding protein that tethers viral episomes to heterochromatic regions of host cell chromosomes and thereby maintains the latent state of the virus. This proposal will test the hypothesis that the regulation of SUMO modification plays a key role in the KSHV life cycle in both establishment of latency and reactivation of this virus by modulating the structure of viral chromatin. In the first Aim, we will investigate the role of K-Rta as a SUMO-dependent ubiquitin ligase in KSHV replication. In the second Aim, the potential mechanisms whereby K-Rta acts in a SUMO-dependent manner and affects the epigenetic signature of viral chromatin will be analyzed. Taken together, these studies will define a new role of K-Rta, as well as LANA, in KSHV replication, and will provide insights into the post- translational modifications and chromatin remodeling that govern latency and reactivation. PUBLIC HEALTH RELEVANCE: The research in this R01 grant application on KSHV will produce a new understanding of regulation of gene expression and epigenetic control. Our research will provide fundamental knowledge regarding the mechanisms of viral reactivation and could produce innovative approaches for the treatment of infectious disease, especially cancer caused by this tumorigenic virus.