GENETICS AND FUNCTIONS OF KSHV GLYCOPROTEINS GM AND GN

KSHV 糖蛋白 GM 和 GN 的遗传学和功能

• 批准号: 7959460
• 负责人: OSWALD D'AUVERGNE
• 金额: $ 9.61万
• 依托单位: LOUISIANA STATE UNIV A&M COL BATON ROUGE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2010-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7959460
• 关键词: B-Cell Lymphomas; Biomedical Research; Body cavities; Cell Line; Cells; Codon Nucleotides; Computer Retrieval of Information on Scientific Projects Database; DEAE-Dextran; Funding; Genes; Glycoproteins; Grant; Human Herpesvirus 8; Infection; Institution; Kaposi Sarcoma; Louisiana; Lymphoma; Lysine; Measurement; Multicentric Angiofollicular Lymphoid Hyperplasia; Mus; Nude Mice; Oncogenic Viruses; Pathogenesis; Pleural effusion disorder; Polybrene; Protamine Sulfate; Protamines; Protein Biosynthesis; Research; Research Personnel; Resources; Role; Small Interfering RNA; Source; System; Time; Transfection; United States National Institutes of Health; Viral; Virion; Virus; Work; base; body cavity; effusion; graduate student; matrigel; polycation; reconstitution; tumor; tumorigenesis

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Kaposi's sarcoma-associated herpesvirus (KSHV), or human herpesvirus 8, is a lymphotropic oncogenic virus that has been implicated in the pathogenesis of Kaposi's sarcoma; body cavity-based B-cell lymphoma (BCBL), or primary effusion lymphoma; and some forms of multicentric Castleman's disease. We have developed a conditional silencing system for partial inhibition of KSHV viral glycoprotein synthesis using specific siRNAs. In this system, protein synthesis can be reconstituted using codon-optimized genes that are not susceptible to siRNA inhibition. We have succesfully used this system to demonstrate that KSHV glycoprotein B (gB) is necessary for virion egress from BCBL-1 cells and virus infectivity (Subramanian, D'Auvergne, et al. J. Virol. 2008, 82:7144-54). To investigate the potential role of gB in tumorigenesis, BCBL-1 cells transiently transfected with anti-gB siRNAs and codon optimized gB were mixed with matrigel and injected subcutaneously in nude mice. Direct measurement of tumors revealed that BCBL-1 cells transfected with anti-gB siRNAs produced tumors significantly smaller than mock-transfected BCBL-1 cells. Co-transfection of codon optimized gB appeared to abrogate the inhibition of tumor formation by siRNAs. These results show that gB is important for infectivity, virion egress and pleural effusion lymphoma (PEL) formation in mice. As part of a graduate student project, we have also worked on the effect of polycations on facilitating KSHV virus entry into 293 cells. It was found that DEAE-dextran, poly-lysine, polybrene, protamine and protamine sulfate enhanced the KSHV entry into 293 and other cell lines when added at the time of infection.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 卡波西肉瘤相关疱疹病毒 (KSHV) 或人疱疹病毒 8 型是一种嗜淋巴细胞致癌病毒，与卡波西肉瘤的发病机制有关。体腔 B 细胞淋巴瘤 (BCBL) 或原发性渗出性淋巴瘤；以及某些形式的多中心卡斯尔曼病。我们开发了一种条件沉默系统，使用特定的 siRNA 部分抑制 KSHV 病毒糖蛋白合成。在该系统中，可以使用不易受 siRNA 抑制影响的密码子优化基因来重建蛋白质合成。我们已成功使用该系统来证明 KSHV 糖蛋白 B (gB) 对于病毒粒子从 BCBL-1 细胞中排出和病毒感染性是必需的（Subramanian, D'Auvergne, et al. J. Virol. 2008, 82:7144-54） 。为了研究gB在肿瘤发生中的潜在作用，将瞬时转染抗gB siRNA和密码子优化的gB的BCBL-1细胞与基质胶混合并皮下注射到裸鼠体内。对肿瘤的直接测量表明，转染抗 gB siRNA 的 BCBL-1 细胞产生的肿瘤明显小于模拟转染的 BCBL-1 细胞。密码子优化的 gB 的共转染似乎消除了 siRNA 对肿瘤形成的抑制。这些结果表明，gB 对于小鼠的感染性、病毒粒子排出和胸腔积液淋巴瘤 (PEL) 形成很重要。作为研究生项目的一部分，我们还研究了聚阳离子对促进 KSHV 病毒进入 293 细胞的影响。结果发现，感染时添加 DEAE-葡聚糖、聚赖氨酸、聚凝胺、鱼精蛋白和硫酸鱼精蛋白可增强 KSHV 进入 293 和其他细胞系的能力。