Functional Specificity of Drosophila homeoprotein DNA binding complexes

果蝇同源蛋白 DNA 结合复合物的功能特异性

• 批准号: 7615884
• 负责人: Zenobia C Cofer
• 金额: $ 4.12万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-03-01 至 2012-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7615884
• 关键词: Abdomen; Anterior; Behavior; Binding; Binding Proteins; Binding Sites; Cell physiology; Complex; DNA Binding; DNA-Binding Proteins; Development; Developmental Process; Diagnosis; Disease; Dissection; Drosophila genus; Ectopic Expression; Electrophoretic Mobility Shift Assay; Elements; Embryo; Enhancers; Family; Gene Expression; Gene Targeting; Genes; Genetic; Genome; Goals; Homeodomain Proteins; Human; In Vitro; Intervention; Knowledge; Lead; Learning; Life; Malignant Neoplasms; Mediating; Methods; Mutation; Nucleotides; Pattern; Process; Protein Binding; Proteins; Reporter; Repression; Role; Site; Specificity; Staging; Tertiary Protein Structure; Testing; Time; Transgenes; Transgenic Organisms; Work; cofactor; gene repression; genetic analysis; in vivo; insight; leukemia; member; mutant; novel strategies; preference; protein expression; public health relevance; research study

DESCRIPTION (provided by applicant): Developmental processes involve regulatory cascades among groups of genes that encode DMA binding proteins, which direct the full range of cellular behaviors required for normal function at all life stages. Much remains to be learned about how members of conserved families of DMA binding proteins act in concert to orchestrate patterns of gene expression. This project will investigate one of the most highly conserved of these families, the homeodomain proteins, which are known to be involved in developmental abnormalities and cancer/leukemia progression, among other important disease processes. The broad, long-term goal of these studies is to understand how DMA binding proteins interact with each other and their target sites in the genome to achieve functional specificity during development. The Engrailed homeodomain protein interacts with several other members of the same DNA binding superfamily to help it find and regulate the appropriate target genes. This proposal is to investigate underlying mechanisms using a combination of in vitro DNA binding analysis and the genetic and transgenic methods available in Drosophila, including reporter transgenes, transgenes that generate ectopic protein expression, and rescuing transgenes that mimic the endogenous pattern of expression, each used in both wildtype and mutant genetic backgrounds. The specific aims are: 1) to identify and dissect binding sites for Engrailed within an identified target enhancer in the direct target gene sloppy-paired through a combination of in vitro binding studies and in vivo transgenic analysis; and 2) to dissect the protein domains necessary for Engrailed to cooperate with its cofactors on those target sites, and test the requirements for those domains for in vivo function. PUBLIC HEALTH RELEVANCE: Fundamental knowledge of mechanisms that underlie cellular processes has proven to be essential for developing truly novel approaches to diagnosis and intervention in disease processes of all kinds, most notably in cancer development, where genetic changes accumulating over time are now understood to result in the abnormal cellular behaviors that lead to cancer. These studies will extend our understanding of how genes regulate other genes by studying a family of interacting DNA binding proteins that are highly conserved from Drosophila to humans.

描述（由申请人提供）：发育过程涉及编码DMA结合蛋白的基因组之间的调节级联，这些基因组指导所有生命阶段正常功能所需的全部细胞行为。关于DMA结合蛋白的保守家族成员如何协同协调基因表达模式，还有很多有待了解的地方。该项目将研究这些家族中最高度保守的同源结构域蛋白之一，已知它参与发育异常和癌症/白血病进展，以及其他重要的疾病过程。这些研究的广泛和长期目标是了解DMA结合蛋白如何相互作用以及它们在基因组中的靶位点，从而在发育过程中实现功能特异性。engriled同源结构域蛋白与同一DNA结合超家族的其他成员相互作用，帮助它找到并调节合适的靶基因。本研究将结合体外DNA结合分析和现有的果蝇遗传和转基因方法，包括报告基因、产生异位蛋白表达的转基因和模拟内源性表达模式的拯救性转基因，在野生型和突变型遗传背景下进行研究。具体目的是：1)通过结合体外结合研究和体内转基因分析，鉴定和解剖直接靶基因马虎配对中已鉴定的靶增强子内Engrailed的结合位点；2)剖析Engrailed与其辅因子在这些靶点上协同工作所必需的蛋白结构域，并测试这些结构域在体内的功能需求。公共卫生相关性：细胞过程背后机制的基本知识已被证明对于开发真正新颖的方法来诊断和干预各种疾病过程至关重要，尤其是在癌症发展中，随着时间的推移，遗传变化的积累现在被理解为导致导致癌症的异常细胞行为。这些研究将通过研究从果蝇到人类高度保守的相互作用的DNA结合蛋白家族，扩展我们对基因如何调节其他基因的理解。