CELLULAR PROLIFERATION AND N-TERMINUS OF SV40 LARGE T ANTIGEN

SV40 大 T 抗原的细胞增殖和 N 末端

• 批准号: 6575613
• 负责人: James A DeCaprio
• 金额: $ 22.84万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-03-01 至 2003-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6575613
• 关键词: cell cycle; cell line; cell proliferation; cell transformation; cyclins; genetic transcription; protein structure function; simian virus 40; temperature sensitive mutant; tumor antigens; tumor promoters; virus antigen

SV40 large T Antigen (T Ag) can serve as both a potent transforming oncoprotein and transcriptional activator. Several domains of T Ag have been identified .that contribute to the transforming and transactivation functions and, in some cases, these domains appear to overlap. However, it is not known whether the ability of T Ag to activate transcription from a wide variety of viral and cellular promoters contributes to cellular transformation. The dual effects of T Ag on the temperature sensitive Syrian hamster cell line, ts 13, may provide the t6ols to address this question. This cell line undergoes a GI arrest at the restrictive temperature due to a mutation in TAFII250, an essential component of the TFIID RNA polymerase Il complex. Transcription from a limited number of promoters including cyclin A is significantly reduced in tsl3 cells at the restrictive temperature. Expression of wild type TAF1I250 or SV40 large T Ag can override the cell cycle block and rescue the cyclin A promoter defects at the restrictive temperature. We propose to perform a genetic and biochemical analysis of the T Ag domains that are required to neutralize the cell cycle and transcriptional defects in tsl3 cells. We will examine whether the ability of T Ag to rescue the cyclin A promoter defect in tsl3 cells correlates with its ability to bind to certain TBP-Associated-Factors (TAFs) contained in the TFIID complex. Finally, we will examine the correlation between T Ag's ability to rescue the cyclin A promoter defect in tsl3 cells with its ability to override a GI/S arrest in these cells at the restrictive temperature. The goal here would be determine whether the specific domains of T Ag required for transactivation and rescue of the cyclin A promoter and for growth promotion in the tsl3 cells at the restrictive temperature were also required for cellular transformation. To the extent that this is so, the information extracted from these experiments could speak to the overall T Ag transforming mechanism.

SV40大T抗原（T Ag）是一种有效的转化癌蛋白和转录激活因子。已经鉴定了T抗原的几个结构域，它们有助于转化和转录激活功能，并且在某些情况下，这些结构域似乎重叠。然而，目前尚不清楚T抗原激活多种病毒和细胞启动子转录的能力是否有助于细胞转化。T抗原对温度敏感的叙利亚仓鼠细胞系ts 13的双重作用可能为解决这一问题提供了t6ols。该细胞系在限制性温度下由于TFII250（TFIID RNA聚合酶II复合物的必要组分）的突变而经历GI停滞。在限制性温度下，从有限数量的启动子（包括细胞周期蛋白A）的转录在tsl3细胞中显著减少。野生型TAF1I250或SV40大T抗原的表达可以克服细胞周期阻滞，并在限制性温度下挽救细胞周期蛋白A启动子缺陷。我们建议进行遗传和生物化学分析的T抗原结构域所需的中和细胞周期和转录缺陷的tsl3细胞。我们将研究T抗原是否有能力拯救细胞周期蛋白A启动子缺陷的tsl3细胞与其能力结合到某些TFIID复合物中所含的TBP相关因子（TAF）。最后，我们将研究T Ag的能力，以挽救在tsl3细胞中的细胞周期蛋白A启动子缺陷与其能力，以覆盖在这些细胞中的GI/S逮捕在限制性温度之间的相关性。这里的目标是确定细胞转化是否也需要细胞周期蛋白A启动子的反式激活和拯救以及在限制性温度下在ts13细胞中促进生长所需的T Ag的特异性结构域。在这种程度上，从这些实验中提取的信息可以说明整个T Ag转化机制。