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USP7 targeting by HHV-8 vIRFs

HHV-8 vIRF 靶向 USP7

基本信息

批准号：
10581544
负责人：
John Nicholas
金额：
$ 40.94万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-03-01 至 2025-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10581544
关键词：
Ablation Acquired Immunodeficiency Syndrome Affect Apoptosis Inhibitor Apoptotic Area B lymphoid malignancy Binding Biological Biology Catalytic Domain Cell Survival Cell physiology Cells Consensus Data Development Disease Endothelium Epithelium Etiology Future Genes Herpesviridae Homologous Gene Human Herpesvirus 8 In Vitro Interferons Kaposi Sarcoma Link Lymphoma cell Lytic Lytic Phase MG132 Maps Mediating Methods Modification Multicentric Angiofollicular Lymphoid Hyperplasia Mutate N-terminal Natural Immunity Pathway interactions Pharmaceutical Preparations Phenotype Polyubiquitin Productivity Proteins Public Health Publishing Refractory Regulation Reporting Research Role Seminal Signal Transduction Stress Substrate Interaction System TRAF Domain Therapeutic Therapeutic Agents Time Transfection Ubiquitin Ubiquitination Variant Viral Viral Proteins Virus Virus Diseases Virus Latency Virus Replication Work Xeroderma Pigmentosum adduct antiviral drug development cell type cellular targeting disorder prevention genetic resistance latency-associated nuclear antigen latent infection lytic replication mutant novel primary effusion lymphoma targeted treatment ubiquitin isopeptidase ubiquitin-specific protease viral interferon regulatory factor viral interferon regulatory factor-3

项目摘要

Human herpesvirus 8 (HHV-8) is involved etiologically in AIDS-associated diseases Kaposi’s sarcoma, primary effusion lymphoma (PEL), and multicentric Castleman’s disease; in all, viral latent and lytic functions are believed to contribute. HHV-8 encodes four interferon regulatory factor homologues (vIRFs) that can interact in inhibitory fashion with cellular IRFs and/or a variety of other cellular proteins involved in innate immunity and antiviral stress signaling. However, very few of these interactions have been detected and assessed functionally in the context of virus infection. It has recently been reported that both vIRF-1 and vIRF-4 interact with the deubiquitinase USP7 [ubiquitin-specific protease 7, also called herpesvirus-associated USP (HAUSP)], and we have identified vIRF-3 (published) and vIRF-2 (new data in this revised application) as additional interaction partners of USP7. While vIRF-4 interacts with the USP7 N-terminal TRAF domain via a core motif, ASTS, matching the consensus USP7-interaction motif, A/PxxS, present in many cellular and viral proteins, vIRF-1 and vIRF-3 have, respectively, a single and duplicated TRAF domain-interacting EGPS core motif and vIRF-2 appears to interact with USP7 in a novel manner. vIRFs 1, 2 and 3 are expressed in latently (in addition to lytically) infected PEL cells, but in other examined cell types, all vIRFs appear to be expressed only during lytic replication. Through depletion-based analyses in PEL cells and gene ablation in the context of HHV-8 bacmid (BAC16) virus-infected iSLK (inducible, epithelial) cells, we have determined that vIRF-1, like vIRF-3, is important for the viability of latently infected PEL cells and that vIRF-3, in contrast to vIRF-1, negatively regulates HHV-8 productive replication. Furthermore, using WT or USP7-refractory (EGPS motif-mutated) vIRF-1/3 “rescue” of vIRF depletion phenotypes in PEL cells and BAC16 mutants expressing USP7-refractory variants of vIRFs 1 and 3, we have determined that vIRF-1 and vIRF-3 interactions with USP7 are involved centrally in the respective latent and lytic activities. USP7 depletion revealed directly the importance of the deubiquitinase for latent PEL cell viability and HHV-8 productive replication. We have also identified ubiquitin modifications of vIRFs 1 and 3, raising the possibility of USP7 modulation of vIRF expression and/or activities via ubiquitin editing. In this application we propose to: (1) examine the functional and biological consequences of vIRF-2 targeting of USP7; (2) characterize structurally and functionally vIRF ubiquitination and USP7 editing thereof; (3) identify vIRF- regulated USP7 targets in the context of HHV-8 infected cells and the functional significance of these proteins in HHV-8 biology. The proposed work will, for the first time, characterize vIRF-2 targeting of USP7 and identify mechanisms underlying biological activities mediated via vIRF-USP7 interactions, of demonstrated importance to both latent and lytic HHV-8 biology. The project will provide mechanistic and biological information (at present sparse) about the significance of viral targeting of USP7 and has the potential to inform future development of novel antiviral and therapeutic agents directed against the mechanisms and pathways identified by this project.

人类疱疹病毒8 （HHV-8）参与艾滋病相关疾病卡波西肉瘤的病因学研究