REGULATION OF THE EBV BRLF1 AND BZLF1 PROMOTERS

EBV BRLF1 和 BZLF1 启动子的监管

• 批准号: 2099487
• 负责人: Shannon Celeste Kenney
• 金额: $ 16.44万
• 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/2099487
• 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EB病毒（EBV）的关键问题 生物学 即刻早期蛋白BZLF1（Z）的过度表达是 足以破坏病毒潜伏期 细胞和病毒调节 因此，这种基因产物的存在对于确定病毒是否 感染是潜伏性的，而不是生产性的。 Z蛋白可以衍生自 两种消息之一：1.0 kb消息（从BZLF 1转录而来 启动子）仅产生Z蛋白，或2.8kb双顺反子信息 （从BRLF1启动子转录），其可以使Z和BRLF1 (R)早期蛋白质 病毒潜伏期的中断 可能是通过激活BZLF1或BZLF2介导的。 BRLF1启动子。 在这项资助中，我们计划鉴定细胞转录因子， 介导病毒潜伏期的破坏，通过激活 BZLF 1或BRLF 1启动子的功能，并确定BZLF 1或BRLF 1启动子的生物学作用。 1.0 kb信息与2.8kb信息作为Z蛋白的来源。 我们 最近发现BZLF1和BRLF1启动子都与 细胞阴阳1（YY-1）蛋白以及oct样蛋白。 我们已经证明，BRLF1（而不是BZLF1）启动子被激活， 由细胞转录因子Sp1控制。 此外，我们还发现， BRLF 1启动子含有zif268和WT 1的结合位点， 转录因子 在我们的第一个具体目标中，我们将确定 与BZLF 1和BRLF 1结合的各种转录因子 启动子，并产生消除这些位点的定点突变。 在 第二个具体目标，我们将审查的功能意义， 与BZLF 1和BRLF 1结合的各种转录因子中的每一种 不同细胞类型的启动子。 在第三个具体目标中，我们将 检查Z和R立即早期蛋白与 显示调节BZLF1和BRLF1启动子的细胞转录因子 功能 在第四个具体目标中，我们将确定 1.0 kb与2.8 kb的信息作为Z蛋白的来源，在各种 EB病毒感染的类型使用PCR分析。 在最后的具体目标中， 将构建突变病毒，其中Z或R蛋白 已被废除，或其中2.8 kb的双顺反子性质 信息已被破坏，并确定生物后果， 这些突变。 这些研究应有助于确定 导致病毒潜伏期的细胞因子以及 BZLF1与BRLF1启动子在破坏病毒潜伏期方面的比较。