BH3-only protein targeting by HHV-8

描述（由申请人提供）：人类疱疹病毒8 （HHV-8）与内皮卡波西肉瘤（KS）和B细胞恶性肿瘤原发性渗出性淋巴瘤和多中心Castleman病相关，所有这些主要发生在HIV合并感染的背景下。潜伏基因和裂解基因产物都被认为是导致这些疾病的原因，而有效复制和病毒载量的增加与KS有关。了解成功裂解复制所需的过程将为靶向基本病毒功能以阻断病毒复制提供机会，从而治疗或预防HHV-8相关疾病。HHV-8和其他病毒复制的关键是限制应激诱导的凋亡信号的病毒机制，这是一种针对病毒感染的先天防御机制。HHV-8编码几种抑制这种促凋亡途径的蛋白质。这些蛋白包括病毒干扰素调节因子-1 （vIRF-1）、病毒G蛋白偶联受体（vGPCR）和病毒趋化因子vCCL-1和vCCL-2，所有这些蛋白都被证明可以抑制细胞凋亡，并且是有效病毒复制所必需的。本实验室已经确定了vIRF-1抑制剂与复制诱导的促凋亡bh3蛋白（BOPs） Bim和Bid的相互作用，并揭示了控制这些蛋白对病毒有效复制的重要性。通过转导的shrna抑制Bim或Bid的表达会导致病毒复制的显著增加，并且破坏vIRF-1:BOP相互作用会抑制病毒的产生

DESCRIPTION (provided by applicant): Human herpesvirus 8 (HHV-8) is associated with endothelial Kaposi's sarcoma (KS) and B cell malignancies primary effusion lymphoma and multicentric Castleman's disease, all of which occur predominantly in the context of HIV coinfection. Both latent and lytic gene products are believed to contribute to these pathologies, and productive replication and increased viral loads are associated with KS. Understanding the processes required for successful lytic replication will provide opportunities for targeting essential viral functions to block virus replication and thereby treat or prevent HHV-8 associated disease. Key to replication by HHV-8 and other viruses are viral mechanisms that limit stress-induced apoptotic signaling that serves as an innate defense mechanism against virus infection. HHV-8 encodes several proteins that inhibit such pro-apoptotic pathways. These proteins include viral interferon regulatory factor-1 (vIRF-1), viral G protein coupled receptor (vGPCR), and viral chemokines vCCL-1 and vCCL-2, all of which have been demonstrated to both inhibit apoptosis and be required for efficient virus replication. This laboratory has identified inhibitor interactions of vIRF-1 with replication-induced pro-apoptotic BH3-only proteins (BOPs) Bim and Bid and revealed the critical importance of controlling these proteins for efficient virus replicaton to occur. Suppression of Bim or Bid expression via transduced shRNAs leads to very significant increases in virus replication, and disruption of vIRF-1:BOP interactions inhibits virus production and promotes apoptosis in lytically infected cells. In addition to Bim and Bid, vIRF-1 targets other BOPs (Bik, Bmf, Hrk, Noxa), via their functional BH3 domains, indicating the biological importance of these BOPs also and the need to inhibit their activities during lytic replication. Furthermore, the viral chemokine receptor (vGPCR) and CCR8 agonists vCCL-1 and vCCL-2 activate signaling pathways that lead to suppression of Bim expression. Thus, chemokine receptor signaling is likely to enhance HHV-8 productive replication in part via BOP regulation. This application proposes to: (1) elucidate the mechanisms of vGPCR and vCCL suppression of Bim, an identified major inhibitor of HHV-8 replication; (2) identify peptide and pharmacological inhibitors of vIRF-1:BOP interactions to specifically inhibit this mechanism of BOP regulation; (3) determine the antiviral activities of lytic cycle-induced BOPs and the individual and combined contributions, via BOP control, of vCCLs, vGPCR, and vIRF-1 to HHV- 8 productive replication. The project comprises a focused analysis of BOP function and targeting by vCCLs, vGPCR and vIRF-1 during HHV-8 lytic replication and assessment of inhibitory reagents and methods that could potentially be developed for antiviral purposes.