COFACTORS AND FUNCTIONS OF HOMEODOMAIN REPRESSORS

描述（由申请人提供）：基因转录的变化在癌症的进展、大多数其他人类疾病、衰老过程以及多细胞生物发展的所有阶段都很重要。详细了解这些变化是如何调节的，是诊断工具和干预策略的基础。进一步的发展有望带来新的方法，并提高现有方法的有效性。重要的是，关于所涉及的基本过程仍然存在许多问题。序列特异性DNA结合蛋白和它们招募到DNA上的辅助因子是最重要的转录调节因子之一。其中最大的一类蛋白质家族共享DNA结合同源结构域基序。在果蝇中可用的工具使得在真实的体内环境中详细研究作用和相互作用的机制成为可能。本研究旨在研究这一家族的几个成员的作用机制，以解决一些基本问题，如如何完成对特定DNA靶点的识别，以及这种识别对抑制转录的辅因子招募的影响，对介导调节的染色质变化的影响，以及对被调节的发育途径的影响。结合体内和体外方法，这些研究将更清楚地了解蛋白质组合如何在体内识别适当的靶位，以及这种识别对下游基因和途径调节的影响。该项目的具体目标是：

DESCRIPTION (provided by applicant): Changes in gene transcription are important in the progression of cancer, in most other human diseases, and in the aging process, as well as in the development of multicellular organisms at all stages. A detailed understanding of how such changes are regulated is the basis of both diagnostic tools and intervention strategies. Further advancement holds the promise of novel approaches, and of increased effectiveness of current approaches. Importantly, many questions remain about the fundamental processes involved. Sequence-specific DNA binding proteins and the cofactors that they recruit to the DNA are among the most important regulators of transcription. One of the largest such families of proteins share the DNA binding homeodomain motif. Tools available in Drosophila make it possible to study mechanisms of action and interaction in detail in a true in vivo context. This proposal is to study mechanisms of action of several members of this family, to address basic questions that remain about how recognition of specific DNA target sites is accomplished, and the consequences of that recognition for the recruitment of cofactors to repress transcription, for chromatin changes that mediate regulation, and for the developmental pathways that are being regulated. Using a combination of in vivo and in vitro approaches, these studies will provide a clearer understanding of how combinations of proteins recognize appropriate target sites in vivo, and the consequences of that recognition for the regulation of downstream genes and pathways. The Specific Aims of the project are: 1) To determine the mechanisms that generate target gene specificity for the homeodomain protein Engrailed. Investigate the mechanisms that lead from DNA binding to repression of the direct target gene sloppy paired. 2) To analyze the functions of an En-interacting protein that contains a zinc-finger DNA binding domain, and to test whether this partner contributes to target gene specificity, or to activation/repression function on specific target sites in vivo. 3) To identify and analyze functional binding sites for the homeodomain-containing represser Evenskipped in the target gene sloppy paired. Determine the mechanisms responsible for the sequential repression during development of sloppy paired by first Even-skipped and then Engrailed.