Gene Activation by Remote Transcriptional Enhancers

描述（来自申请人摘要的逐字）：激活 启动子几个碱基的距离是至关重要的适当表达 后生动物发育所需的几个基因。我们发现了 果蝇Nipped-B蛋白在遗传筛选中支持 被切割基因中的远距离增强子激活。Nipped-B也有助于激活 的Ultrabithorax（Ubx）基因的远程增强子，并参与姐妹 分裂的幼虫神经母细胞中的染色单体凝聚。我们的目标是了解 Nipped-B如何参与基因激活Nipped-B与人类同源 功能未知的蛋白质和酵母Scc 2（S.酿酒酵母（S. cerevisiae）和Mis 4（S. 粟酒裂殖酵母）姐妹染色单体凝聚所需的蛋白质。Scc 2是需要的， 一种将姐妹染色单体结合在一起以与 染色体数据表明Scc 2和Mis 4完全与染色体结合， 细胞周期的各个阶段，并帮助在S期招募粘附素。的 研究人员推测，Nipped-B的作用类似于帮助基因激活， 募集作用于染色体结构的蛋白质复合物， 和发起人紧密联系在一起。黏连蛋白含有两种SMC蛋白， 直接影响DNA和染色体结构。在拟议的工作中，突变 在编码SMC蛋白质或其它粘着蛋白复合物组分的基因中， 用于确定这些蛋白质是否也参与cut或 ubx作为一种更普遍的方法，与Nipped-B相互作用的蛋白质将被 分离的生化，然后突变的基因编码他们将是 用于探索它们在基因表达和染色单体凝聚中的体内作用。 将使用Nipped-B中的突变和构建的缺失突变体来观察是否 Nipped-B蛋白的基因表达和染色单体凝聚功能 是可分离的。免疫染色和染色质免疫沉淀实验 将用于确定Nipped-B蛋白是否直接作用于cut和Ubx，或 在远离这些靶基因的位点。这些实验将有助于定义 Nipped-B在基因激活中作用，并增加了对Nipped-B如何激活的理解 远程增强子调节转录。这将有助于阐明 一些人类基因表达缺陷的潜在机制 疾病

DESCRIPTION (Verbatim from the applicant's abstract): Enhancers that activate promoters several kilobases away are vital for the appropriate expression of several genes that are required for metazoan development. We discovered the Drosophila Nipped-B protein in a genetic screen for factors that support activation by a distant enhancer in the cut gene. Nipped-B also aids activation of the Ultrabithorax (Ubx) gene by remote enhancers, and participates in sister chromatid cohesion in dividing larval neuroblasts. The goal is to understand how Nipped-B participates in gene activation. Nipped-B is homologous to a human protein of unknown function and to the yeast Scc2 (S. cerevisiae) and Mis4 (S. pombe) proteins required for sister chromatid cohesion. Scc2 is needed for the Cohesin protein complex that holds sister chromatids together to bind to chromosomes. The data indicate that Scc2 and Mis4 bind to chromosomes at all stages of the cell cycle and help recruit Cohesin during S phase. The investigator postulates that Nipped-B acts similarly to aid gene activation by recruiting protein complexes that act on chromosome structure to hold enhancers and promoters close together. Cohesin contains two SMC proteins, which may directly affect DNA and chromosome structure. In the proposed work, mutations in genes encoding SMC proteins or other components of the Cohesin complex will be used to determine if these proteins also participate in activation of cut or Ubx. As a more general approach, proteins that interact with Nipped-B will be isolated biochemically, and then mutations in the genes encoding them will be used to explore their in vivo roles in gene expression and chromatid cohesion. Mutations in Nipped-B and constructed deletion mutants will be used to see if the gene expression and chromatid cohesion functions of the Nipped-B protein are separable. Immunostaining and chromatin immunoprecipitation experiments will be used to determine if Nipped-B protein acts directly at cut and Ubx, or at sites distant from these target genes. These experiments will help define the role of Nipped-B in gene activation and increase understanding of how remote enhancers regulate transcription. This will help illuminate the basic mechanisms underlying gene expression defects that occur in some human diseases.