E2F-6 and Repression of p19ARF Gene Expression

E2F-6 和 p19ARF 基因表达的抑制

• 批准号: 6615050
• 负责人: NICHOLAS H HEINTZ
• 金额: $ 0.06万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-15 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6615050
• 关键词: DNA replication origin; apoptosis; artificial chromosomes; binding sites; cell cycle; electroporation; fluorescence microscopy; gene expression; gene induction /repression; genetic regulatory element; genetic transcription; genetically modified animals; laboratory mouse; messenger RNA; microarray technology; northern blottings; polymerase chain reaction; southern blotting; tissue /cell culture; transcription factor; transfection

DESCRIPTION (provided by applicant) The unusual organization of INK4a/ARF gene locus permits the expression of two overlapping transcripts that are translated in alternate reading frames. Recently we have developed a novel and highly efficient method for the transfer of bacterial artificial chromosomes (BACs) into cells in culture. Combined with the ability to introduce specific modifications in BACs by homologous recombination, this novel gene transfer system provides an opportunity to dissect the role of specific regulatory elements in the context of complex genetic loci, both in vitro and in BAC transgenic mice. Using this gene transfer system and modified alleles of INK4a/ARF, we propose to: 1) investigate the function of a TTTCCCGC E2F binding site in activation of pl9ARF mRNA expression in response to the transcriptional activators E2Fs 1-3, and 2) examine the specific role of E2F-6 in repression of pl9?ARF transcription. These studies will develop a new paradigm for dissecting the role of specific regulatory elements within the context of entire genetic loci in cells in culture, thereby permitting evaluation of specific BAC alleles prior to the generation of BAC transgenic mice.

描述（由申请人提供） INK 4a/ARF基因位点的不寻常组织允许表达两个 在交替阅读框架中翻译的重叠转录本。 最近，我们开发了一种新的和高效的方法， 将细菌人工染色体（BAC）植入培养细胞中。结合 在BAC中引入特异性修饰的能力， 重组，这种新的基因转移系统提供了一个机会， 在复杂的环境中， 遗传位点，无论是在体外和BAC转基因小鼠。利用这个基因 INK 4a/ARF的转移系统和修饰的等位基因，我们建议：1） 研究TTTCCCGC E2 F结合位点在p19 ARF活化中的功能 响应于转录激活因子E2 Fs 1-3的mRNA表达，和2） 研究E2 F-6在p19抑制中的具体作用？ARF转录。 这些研究将发展一种新的范式， 在细胞中的整个遗传基因座的背景下的调节元件， 培养，从而允许在培养前评估特定的BAC等位基因。 BAC转基因小鼠的产生。