CARD14 is essential for hepatitis C virus replication and activation of NFKB

描述(申请人提供)：丙型肝炎病毒(HCV)感染困扰着全球约1.7亿人，这些人中的慢性病毒感染是导致肝细胞癌(HCC)的重要原因。丙型肝炎病毒诱导肝细胞癌的机制目前尚不清楚，但很可能是病毒引起宿主细胞信号通路的改变，从而为病毒创造一个复制生态位，避免宿主的抗病毒程序是其发病机制的重要组成部分。利用基因组规模的siRNA筛选，我们已经确定宿主蛋白CARD14是丙型肝炎病毒RNA复制的重要因素。CARD14是已知的核因子-kB途径的调节因子，但其在生物学中的确切功能尚不清楚。我们的初步数据显示，丙型肝炎病毒感染细胞导致核因子-kB活性急剧增加，这种激活依赖于CARD14蛋白的存在。许多感染性因素通过改变NF-kB基因的表达来促进宿主细胞的存活，而在慢性丙型肝炎病毒感染过程中对这一途径的长期操纵可能是导致癌症发生的因素之一。我们的假设是，依赖于CARD14的核因子-kB通路的激活是有效的丙型肝炎病毒RNA复制和防止感染细胞中的凋亡所必需的，并且丙型肝炎病毒对这一通路的操纵是肝细胞癌发生发展的一个重要因素。关于CARD14如何在NF-kB信号转导中发挥作用，病毒如何操纵这一途径，以及这种操纵导致基因表达发生什么变化的细节是需要解决的重要问题，我们设计了四个具体目标来解决这些重要问题。我们建议验证CARD14和NF-kB激活之间的联系，绘制CARD14的哪些区域是NF-kB激活和丙型肝炎病毒复制所必需的，确定CARD14是否作为细胞凋亡的调节因子，定义CARD14信号通路的一部分，并确定CARD14在丙型肝炎病毒感染过程中操纵哪些细胞基因。这些目标的完成将大大增加我们对丙型肝炎病毒复制的细胞需求，丙型肝炎病毒操纵核因子-kB信号以促进细胞生存，以及CARD14在这些过程中的功能的理解。这些实验对丙型肝炎病毒感染者的肝细胞癌的发展具有重要的潜在意义。 公共卫生相关性：该项目涉及丙型肝炎病毒与一种名为CARD14的细胞蛋白的相互作用。CARD14参与调节细胞生存和生长的重要细胞途径。丙型肝炎病毒和CARD14的相互作用可能对丙型肝炎感染患者的肝细胞癌的发展具有重要意义。

DESCRIPTION (provided by applicant): Hepatitis C virus (HCV) infection afflicts approximately 170 million people worldwide and chronic virus infection in these individuals is a significant cause of hepatocellular carcinoma (HCC). The mechanisms by which HCV induces HCC are largely unknown, but it is likely that virus induced alteration of host cell signaling pathways to create a replication niche for the virus and avoid the host anti-viral program is an important component of this pathogenesis. Using genomic scale siRNA screening, we have identified the host protein CARD14 as an important factor for HCV RNA replication. CARD14 is a known regulator of the NF-kB pathway, but its precise function in biology is unknown. Our preliminary data shows that infection of cells by HCV leads to a dramatic increase in NF-kB activation, and this activation is dependent on the presence of the CARD14 protein. Many infectious agents alter NF-kB gene expression to promote survival of the host cell, and the long-term manipulation of this pathway during chronic HCV infection may be one factor leading to the development of cancer. Our hypothesis is that CARD14 dependent activation of the NF-kB pathway is required for efficient HCV RNA replication and the prevention of apoptosis in infected cells, and that manipulation of this pathway by HCV is an important factor in the development of HCC. The details of how CARD14 functions in NF-kB signaling, how the virus manipulates this pathway, and what changes in gene expression result from this manipulation are important questions that need to be addressed, and we have designed four specific aims to address these important questions. We propose to validate the connection between CARD14 and NF-kB activation, map what regions of CARD14 are required for NF-kB activation and HCV replication, determine if CARD14 is functioning as a regulator of apoptosis, define proteins that are part of the CARD14 signaling pathway, and determine what cellular genes CARD14 manipulates during HCV infection. The completion of these aims will add considerably to our understanding of the cellular requirements for HCV replication, the manipulation of NF-kB signaling by HCV to promote cell survival, and the function of CARD14 in these processes. These experiments are of great potential significance to the development of HCC in those infected with HCV. PUBLIC HEALTH RELEVANCE: This project involves the interaction of the hepatitis C virus with a cellular protein called CARD14. CARD14 is involved in an important cellular pathway regulating cell survival and growth. The interaction of HCV and CARD14 may have significance to the development of hepatocellular carcinoma; a form of liver cancer, in hepatitis C infected patients.