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） INK4A/ARF基因基因座的不寻常组织允许两个表达 重叠的成绩单以替代阅读帧翻译。 最近，我们开发了一种新颖且高效的转移方法 细菌人造染色体（BAC）培养细胞。与 通过同源物引入BAC中特定修改的能力 重组，这个新颖的基因转移系统为 在复杂的背景下剖析特定调节元素的作用 遗传基因座，无论是体外还是在BAC转基因小鼠中。使用此基因 转移系统和Ink4a/arf的修改等位基因，我们建议：1） 研究TTTCCCGC E2F结合位点在PL9ARF激活中的功能 响应转录激活剂E2FS 1-3和2的mRNA表达） 检查E2F-6在抑制PL9 arf转录中的特定作用。 这些研究将开发出一种新的范式来剖析特定的作用 在细胞中整个遗传基因座的背景下的调节元素 培养，从而允许评估特定的BAC等位基因 BAC转基因小鼠的产生。