E2F-6 and Repression of p19ARF Gene Expression

描述(由申请人提供) INK4a/ARF基因座的异常组织允许两个 在交替阅读框架中翻译的重叠的抄本。 最近，我们开发了一种新颖而高效的转移方法 将细菌人工染色体(BAC)植入培养细胞中。与 通过同源基因在BAC中引入特定修饰的能力 重组，这种新的基因转移系统提供了一个机会 剖析特定监管要素在复杂背景下的作用 在体外和在BAC转基因小鼠中的遗传位点。利用这种基因 INK4a/ARF的转移系统和修饰的等位基因，我们建议：1) TTTCCCGC E2F结合位点在pl9ARF激活中的作用 转录激活剂E2Fs1-3和2的mRNA表达 检测E2F-6在抑制PL9？ARF转录中的特异性作用。 这些研究将开发一种新的范式来剖析特定的 细胞中整个遗传基因座背景下的调控元件 培养，从而允许在检测之前评估特定的BAC等位基因 BAC转基因小鼠的产生。

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.