HMG-1 AND HMG-2 AND EUKARYOTIC TRANSCRIPTION

HMG-1 和 HMG-2 以及真核转录

• 批准号: 2603478
• 负责人: WILLIAM M SCOVELL
• 金额: $ 9.65万
• 依托单位: BOWLING GREEN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-07-01 至 2002-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2603478
• 关键词: DNA directed RNA polymerase; SDS polyacrylamide gel electrophoresis; chimeric proteins; genetic promoter element; genetic transcription; immunoprecipitation; oncoproteins; protein binding; protein purification; protein sequence; retinoblastoma protein; transcription factor; tumor suppressor proteins; western blottings

DESCRIPTION: The investigators would like to extend their understanding of the mechanisms by which HMG-1 protein participates in the regulation of gene expression by RNA pol II. HMG-1 represses basal level transcription, while enhancing the level of transcription activated by Oct-1 and USF. They will determine the nature of the HMG-1 interactions with general transcription factors, TBP and TFIIB; activator oncoprotein E1A; and the products of tumor suppressor genes, Rb and p53. The binding will be examined both in the presence and in the absence of a DNA promoter. Of particular interest will be the competitive interactions which the HMG proteins engage in with other transcription factors in the process of exerting their influence to control gene expression and the regulation of the cell cycle. Protein-protein interactions of HMG-1 with regulatory proteins will be examined using GST-fusion capture experiments and analyzed by SDS-PAGE and western blots. A major aim will be to provide an experimental basis which begins to explain the apparent dichotomy that HMG-1 serves as a repressor for basal level transcription, while acting to enhance activated transcription in other characterized promoter systems. HMG-1 binds to the TBP/TATA complex to produce a transcription "dead end" complex, which we believe may be competed off by activators. In addition, the occurrence of protein-protein interactions in solution may play an important, but indirect role. HMG-1 may interact with an activator in increase its effectiveness so that activators would interact more effectively with GTFs and stimulate the level of transcription. The activator proteins may then serve both as an anti-repressor and activator, while HMG-1 would serve as either a repressor and/or coactivator, depending on the circumstance. The investigators shall use findings with E1A and p53 as a tentative model to compare with. On the other hand, HMG-1 may serve as a cellular activator by competing with the E2F regulator transcription factor to stimulate the dissociation of the repressor, Rb, from the Rb/E2F complex. HMG-1 has the minimal recognition sequence for binding to the "A/B box" in the Rb protein. They will map the region(s) of HMG-1 and Rb involved in the direct interaction. By its complexation with both TBP and Rb, HMG-1 may play a pivotal "linking role" in the concerted regulatory action between transcriptional regulation, the control of the cell cycle, cell proliferation and the development of cancer. It is important to note in this respect that both HMG-1 and E2F bind to both TBP and Rb and that the interaction of E2F and Rb are mutually exclusive. The finding from the proposed studies shall provide additional new insights into the possible mechanisms of action for HMG-1 protein in the regulation of gene expression in normal and unregulated cells.

描述：调查人员想扩展他们的理解 HMG-1蛋白参与调节的机制 RNA Pol II的基因表达。 HMG-1压抑基础水平 转录，同时增强了被激活的转录水平 Oct-1和USF。 他们将确定HMG-1相互作用的性质 与一般转录因子，TBP和TFIIB；激活剂癌蛋白 e1a;以及肿瘤抑制基因RB和p53的产物。 这 在存在和不存在的情况下，将检查结合 DNA启动子。 特别感兴趣的将是竞争性 HMG蛋白与其他转录进行的相互作用 在控制基因的影响的过程中 表达和细胞周期的调节。 蛋白质蛋白质 HMG-1与调节蛋白的相互作用将使用 GST融合捕获实验并通过SDS-PAGE和WESTER进行分析 印迹。 主要目的是提供一个实验基础 解释HMG-1作为阻遏物的明显二分法 基础水平的转录，同时发挥激活 在其他特征启动子系统中的转录。 HMG-1与 TBP/TATA复合物可产生转录“死胡同”复合物， 我们认为这可能会由激活剂竞争。 另外， 溶液中蛋白质 - 蛋白质相互作用的发生可能会发挥 重要但间接的角色。 HMG-1可能与激活剂相互作用 提高其有效性，以使激活因素可以交互更多 有效地使用GTF并刺激转录水平。 这 然后，激活蛋白可以既可以用作抗抑郁药，又可以用作 激活剂，而HMG-1将用作阻遏物和/或 共激活因子，具体取决于情况。 调查人员应使用 具有E1A和p53的发现作为暂定模型。 在 另一方面，HMG-1可以通过与 E2F调节剂转录因子刺激解离 RB/E2F复合物的阻遏物RB。 HMG-1的最小 在RB蛋白中与“ A/B盒”结合的识别顺序。 他们将绘制直接涉及的HMG-1和RB的区域 相互作用。 通过与TBP和RB的络合，HMG-1可以玩 在一致的监管行动中，关键的“链接角色” 转录调控，细胞周期的控制，细胞 增殖和癌症的发展。重要的是要注意 HMG-1和E2F都结合了这两个方面 TBP和RB，E2F和RB的相互作用是相互的 独家的。 拟议研究的发现应提供额外的新 洞悉HMG-1蛋白在 调节正常和不调节细胞中基因表达。