Functional Anatomy of Mammalian Phosphatidylinositol Transfer Proteins

 描述(申请人提供)：这项研究的目标是对一类未被研究的蛋白质进行详细的分析：哺乳动物的磷脂酰肌醇/磷脂酰胆碱转移蛋白(PITPs)。哺乳动物PITP的功能和机制仍有待阐明。该研究计划旨在确定特定哺乳动物PITP亚型--第一类PITP的功能机制。这一建议是基于我们创建和鉴定了PITP基因敲除小鼠和该蛋白功能分级缺陷的一系列等位基因小鼠。Pitp0/0 老鼠及其衍生物是有吸引力的疾病模型，因为它们出生时是活着的，但出生后表现出强大的表型。这些表型允许收集已定义的基于细胞的试剂进行分析。使用这些独特的动物和细胞模型作为主要的分析对象，我们将进行三条线的调查。首先，我们将使用复杂的生化和生物物理方法来破译1类PITPs如何在单分子水平上发挥作用。其次，我们将使用复杂和定量的成像和分析方法来确定1类PITPs如何在功能上与表皮生长因子受体和血小板衍生生长因子受体等信号受体酪氨酸激酶结合。这些受体的适当活性是适当的细胞生长控制和形态反应所必需的。精神错乱会导致癌症和神经缺陷。第三，我们将开发我们组装的强大的筛选平台，以识别和验证具有高特异性的1类PITPs的小分子抑制剂。这些试剂将作为外科手术操作细胞内肌醇磷脂信号的工具化合物具有价值，并有可能识别用于治疗癌症或神经缺陷的新药的先导化合物。PITPs在调节与不同细胞过程相互作用的信号转导通路中发挥核心作用。然而，潜在的机制还不清楚，因为这些都还没有被研究过。Bankaitis实验室是解决1类PITP功能机制问题的唯一准备，因为它开发了独特的实验系统用于分析。

 DESCRIPTION (provided by applicant): The objective of this research is to undertake a detailed analysis of an under-investigated class of proteins: the mammalian phosphatidylinositol/phosphatidylcholine transfer proteins (PITPs). The functions and mechanisms of mammalian PITP function remains to be elucidated. The research plan is designed to identify mechanisms of function of specific mammalian PITP isoforms - the Class 1 PITPs. This proposal is founded on our creation and characterization of a PITP knockout mouse and an allelic series of mice with graded defects in function of this protein. The pitp0/0 mouse and its derivatives are attractive disease models in that these are born alive, but manifest powerful phenotypes after birth. These phenotypes permit collection of defined cell-based reagents for analysis. Using these unique animal and cellular models as primary analytical subjects, we will undertake three lines of investigation. First, we will use sophisticaed biochemical and biophysical approaches to decipher how Class 1 PITPs function at the level of single molecules. Second, we will use sophisticated and quantitative imaging and analytical assays to determine how Class 1 PITPs functionally engages signaling receptor tyrosine kinases such as epidermal growth factor receptor and platelet-derived growth factor receptor. The appropriate activity of these receptors is required for proper cell growth control and morphogen responses. Derangements lead to cancer and neurological deficits. Third, we will exploit a powerful screening platform we assembled for the purpose of identifying and validating small molecule inhibitors with high specificity for Class 1 PITPs. Such reagents would be of value as tool compounds for surgical manipulation of phosphoinositide signaling in cells, and hold the potential of identifying lead compounds for new pharmaceuticals directed at treatment of cancer or neurological deficits. PITPs play central roles in regulating signal transduction pathways that interface with diverse cellular processes. Yet, the underlying mechanisms are not understood because these have not been investigated. The Bankaitis laboratory is uniquely poised to address questions of mechanism of Class 1 PITP function as it has developed unique experimental systems for analysis.