REGULATION OF THE EBV BRLF1 AND BZLF1 PROMOTERS

EBV BRLF1 和 BZLF1 启动子的监管

• 批准号: 2099486
• 负责人: Shannon Celeste Kenney
• 金额: $ 15.49万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-08-01 至 1998-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2099486
• 关键词: B lymphocyte; Burkitt's lymphoma; DNA footprinting; DNA methylation; Epstein Barr virus; chemical binding; epithelium; gel mobility shift assay; gene induction /repression; genetic promoter element; latent virus infection; molecular site; polymerase chain reaction; site directed mutagenesis; tissue /cell culture; transcription factor; virus genetics; virus protein

The control of viral latency is a key issue in Epstein-Barr virus (EBV) biology. Overexpression of the immediate-early protein, BZLF1 (Z), is sufficient to disrupt viral latency. the cellular and viral regulation of this gene product is therefore crucial in determining whether virus infection is latent versus productive. The Z protein can be derived from either of two messages: a 1.0 kb message (transcribed from the BZLF1 promoter) making only Z protein, or a 2.8 kb bicistronic message (transcribed from the BRLF1 promoter) which can make both the Z and BRLF1 (R) immediate-early proteins. Disruption of viral latency could potentially be mediated through activation of either the BZLF1 or the BRLF1 promoter. In this grant we propose to identify the cellular transcription factors mediating disruption of viral latency through activation of either the BZLF1 or the BRLF1 promoter and to determine the biological role of the 1.0 kb message versus the 2.8 kb message as a source of Z protein. We have recently found that both the BZLF1 and BRLF1 promoters are bound by the cellular Yin Yang 1 (YY-1) protein as well as an oct-like protein. We have shown that the BRLF1 (but not the BZLF1) promoter is activated by the cellular transcription factor, Sp1. In addition, we have found that the BRLF1 promoter contains binding sites for the zif268 and WT1 transcription factors. In our first specific aim we will determine the exact binding sites of the various transcription factors binding to the BZLF1 and BRLF1 promoters and create site-directed mutations abolishing these sites. In the second specific aim we will examine the functional significance of each of the various transcription factors binding to the BZLF1 and BRLF1 promoters in different cell types. In the third specific aim we will examine the interaction of the Z and R immediate-early proteins with the cellular transcription factors shown to regulate BZLF1 and BRLF1 promoter function. In the fourth specific aim we will determine the role of the 1.0 kb versus the 2.8 kb message as a source of Z protein in various types of EBV infection using PCR analysis. In the final specific aim we will construct mutant viruses in which either the Z or the R proteins have been abolished, or in which the bicistronic nature of the 2.8 kb message has been destroyed, and determine the biological consequences of these mutations. These studies should help to establish the nature of the cellular factors contributing to viral latency and the role of the BZLF1 versus the BRLF1 promoter in the disruption of viral latency.

病毒潜伏期的控制是爱泼斯坦 - 巴尔病毒（EBV）的关键问题 生物学。 直接蛋白质BZLF1（z）的过表达为 足以破坏病毒潜伏期。 细胞和病毒调节 因此，该基因产物的生产对于确定病毒是否至关重要 感染是潜在的与生产性的。 Z蛋白可以从 两条消息之一：1.0 kb消息（从BZLF1转录 启动子）仅制作Z蛋白，或2.8 kb的双散发信息 （从BRLF1启动子转录）可以同时使Z和BRLF1同时制造 （r）立即蛋白质。 病毒潜伏期的破坏可能 可能通过激活BZLF1或 BRLF1启动子。 在这笔赠款中，我们建议确定细胞转录因子 通过激活任何一种 BZLF1或BRLF1启动子，并确定 1.0 kb消息与2.8 kb消息作为Z蛋白的来源。 我们 最近发现BZLF1和BRLF1启动子都受 细胞阳1（YY-1）蛋白以及OCT样蛋白。 我们已经表明，BRLF1（而非BZLF1）启动子被激活 通过细胞转录因子SP1。 此外，我们发现了 BRLF1启动子包含ZIF268和WT1的结合位点 转录因子。 在我们的第一个特定目标中，我们将确定 与BZLF1和BRLF1结合的各种转录因子 启动子并创建以站点为导向的突变，以消除这些地点。 在 第二个特定目的我们将研究的功能意义 与BZLF1和BRLF1结合的各种转录因子中的每一个 不同细胞类型的启动子。 在第三个特定目标中，我们将 检查Z和R即时蛋白质与该蛋白与该蛋白的相互作用 显示调节BZLF1和BRLF1启动子的细胞转录因子 功能。 在第四个特定目标中，我们将确定 1.0 kb与2.8 kb消息作为各种Z蛋白的来源 使用PCR分析的EBV感染类型。 在最终的特定目的中 将构建Z或R蛋白的突变病毒 被废除了，或者是2.8 kb的双散性性质 信息已被破坏，并确定 这些突变。 这些研究应有助于确定 导致病毒潜伏期和作用的细胞因素 BZLF1与BRLF1启动子在破坏病毒潜伏期中。