Retinal Patterning: A role for protein kinase CK2

视网膜图案：蛋白激酶 CK2 的作用

• 批准号: 7486814
• 负责人: ASHOK BIDWAI
• 金额: $ 28.71万
• 依托单位: WEST VIRGINIA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-01 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7486814
• 关键词: Adult; Alleles; BHLH Protein; Binding; Binding Sites; C-terminal; Cells; Complex; DNA Binding; Defect; Development; Drosophila genus; Enhancers; Eye; Eye Development; Family; Future; Gene Proteins; Gene Targeting; Genes; Grant; Homo; Homologous Gene; Lateral; Mediating; Mediator of activation protein; Modeling; Modification; Molecular; Molecular Conformation; Mus; Mutation; Neurons; Notch Signaling Pathway; Numbers; Optic vesicle; Oranges; Organ; Pattern; Phenotype; Phosphorylation; Phosphorylation Inhibition; Photoreceptors; Principal Investigator; Progress Reports; Proteins; Regulation; Repression; Resolution; Retina; Retinal; Retinal Defect; Role; Role playing therapy; Sensory; Signal Pathway; Signal Transduction; Site; System; Temperature; Testing; Transcription Coactivator; Variant; Work; base; casein kinase II; cell type; conformational conversion; dimer; dosage; fly; histogenesis; imaginal disc; in vivo; insight; member; notch protein; novel; programs; relating to nervous system

DESCRIPTION (provided by applicant): Retinal patterning involves the specification of a diverse array of cell fates through a complex interplay of signaling pathways, using molecular factors and regulatory mechanisms that appear highly conserved. One of these is the Notch signaling pathway. It plays roles during the specification of photoreceptors such as the 'founding' R8's, the R3/4 pairs and the R7 cells. Notch mediates these effects by regulating the expression of a family of structurally related basic-helix-loop-helix (bHLH) proteins encoded by the Enhancer of Split Complex, E(spl)C. These bHLH proteins inhibit the functions of other genes/proteins whose activities, in turn, are required for specification of these neuronal cell fates. Because the inhibition by E(spl) proteins of the proneural genes is central to retinal patterning, substantial efforts have been devoted to unraveling the underlying mechanisms. Our work is focused on the mechanism by which protein kinase CK2 contributes to the regulation of E(spl) proteins via phosphorylation. Our analysis of this modification has uncovered a novel aspect of regulation, whereby phosphorylation of E(spl) appears to mediate a conformational change that is required for repression. Through these studies, CK2 is implicated as a mediator of inhibitory Notch signaling. Our studies will continue to exploit the broad potential of the Drosophila system in order to extend our characterization of the regulation of E(spl) by CK2 during retinal patterning. We now propose three aims: (1) We will analyze variants of E(spl)M8 to identify the in vivo roles of secondary phosphorylation and the mechanism by which this modification may contribute to conformational transitions. (2) We will analyze the mechanistic connection between CK2 and E(spl) using the retinal defects of Nspl, a well described and widely studied Notch mutation. (3) Using conditional (temperature sensitive) mutations in CK2, we will analyze its roles during the R3/4 and R7 cell fates and conduct screens to identify eye-specific factors that regulate repression by E(spl). Our studies are likely to be relevant to mammalian eye development because CK2 regulates, in a remarkably similar manner, the functions of mouse Hes6, a homolog of fly E(spl) proteins. Our studies will clarify the roles of CK2 and may provide insights into how misregulated cell fate specification contributes to eye defects. A successful resolution of these issues will expose a layer of complexity of Notch signaling previously unknown and unexplored.

描述（由申请人提供）：视网膜模式涉及通过使用分子因子和似乎高度保守的调节机制来通过信号通路的复杂相互作用来规范各种细胞命运。其中之一是Notch信号通路。它在诸如“启动” R8，R3/4对和R7细胞等光感受器的规范中扮演角色。 Notch通过调节结构相关的基本螺旋 - 环螺旋（BHLH）蛋白的表达来介导这些效果，该蛋白由Split Complecter的增强剂E（SPL）c编码。这些BHLH蛋白抑制了其他基因/蛋白的功能，而这些基因/蛋白的活性反过来又是这些神经元细胞命运所必需的。由于胸膜基因的E（SPL）蛋白的抑制是视网膜模式的核心，因此已致力于揭示基本机制。我们的工作集中于蛋白激酶CK2通过磷酸化对E（SPL）蛋白的调节的机制。我们对这种修饰的分析发现了调节的新方面，从而使E（SPL）的磷酸化似乎介导了抑制所需的构象变化。通过这些研究，CK2被视为抑制性缺口信号传导的介体。我们的研究将继续利用果蝇系统的广泛潜力，以扩展我们在视网膜模式期间通过CK2对E（SPL）调节的表征。我们现在提出三个目标：（1）我们将分析E（SPL）M8的变体，以识别次级磷酸化的体内作用以及这种修饰可能有助于构象转变的机制。 （2）我们将使用NSPL的视网膜缺陷分析CK2和E（SPL）之间的机械连接，NSPL是一个很好的描述且广泛研究的Notch突变。 （3）使用CK2中的条件（温度敏感）突变，我们将在R3/4和R7细胞命运中分析其作用，并进行筛选，以识别e（SPL）调节抑制作用的眼睛特异性因素。我们的研究可能与哺乳动物的眼睛发育有关，因为CK2以非常相似的方式调节小鼠hes6的功能，这是蝇E（SPL）蛋白的同源物。我们的研究将阐明CK2的作用，并可以提供有关细胞命运规范如何导致眼部缺陷的见解。这些问题的成功解决将揭示先前未知且未探索的Notch信号的复杂性。