Ptdins Transfer Protein Function and Neurodegeneration

描述(由申请人提供)： 这项研究的目的是对一类研究不足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)。PITPs在哺乳动物细胞中的功能和作用机制尚不清楚。该研究计划旨在确定特定哺乳动物PITP亚型(PITPalpha)的功能机制。这一建议是建立在我们创造和表征PITPalpha基因敲除小鼠的基础上的。PITP-/-小鼠患有脊髓-小脑变性、低血糖，以及无法通过小肠吸收膳食脂肪和脂溶维生素(即乳糜粒滞留疾病)。这些表型在出生时就会显现出来，PITPalpha-/-小鼠很快就会屈服于这些疾病。PITPalpha-/-小鼠是一种理想的疾病模型，因为它是活着出生的，但表现出强大的表型。表型出现的速度很快。使用这一独特的模式，我们将进行三条线的调查。 首先，我们将恢复PITPalpha在PITPalpha-/-小鼠的小肠中的表达，以验证我们的假设，即饮食脂肪吸收不良综合征是低血糖和脊髓小脑变性的主要因素。在这项工作中，我们建议建立一种培养的肠细胞模型，在该模型中可以详细研究乳糜粒的组装和运输。其次，我们的数据表明，PITPalpha缺陷的神经元表现出内在的缺陷；最有可能的是通过所谓的‘生存通路’发出信号。使用遗传、生化和信号传递方法，我们将在PITPalpha-/-模型中检验这一假说。这些研究将通过开发一种新的果蝇系统来研究PITPct信号功能来补充。第三，我们将使用基因组工程方法从功能上剖析单个PITPalpha磷脂转移活动的生理功能。特别是，我们将测试PITPalpha的关键生理功能是否促进肌醇磷脂的合成，如果是的话，哪些特定的肌醇磷脂是以依赖于PITPalpha的方式调节的，以及涉及哪些信号通路。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。我们现在确定这些功能界面与人类乳糜粒滞留疾病、葡萄糖稳态失调和脊髓小脑退行性变有关。Bankaitis实验室独一无二地准备解决PITPalpha功能的机制问题，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 function of PITPs in mammalian cells remain to be elucidated. The research plan is designed to identify mechanisms of function of a specific mammalian PITP isoform (PITPalpha). This proposal is founded on our creation and characterization of a PITPalpha knockout mouse. PITP-/- mice suffer from spinocerebellar degeneration, hypoglycemia, and failure to absorb dietary fat and fat-soluble vitamins across the small intestine (i.e. a chylomicron retention disease). These phenotypes manifest themselves upon birth and PITPalpha-/- mice rapidly succumb to these disorders. The PITPalpha-/- mouse is an ideal disease model in that it is born alive, but manifests powerful phenotypes. The rates of phenotype onset are rapid. Using this unique model, we will undertake three lines of investigation. First, we will restore PITPalpha expression to the small intestine of the PITPalpha-/- mouse to test our hypothesis that the dietary fat malabsorption syndrome is a primary factor in hypoglycemia and spinocerebellar degeneration. In this effort, we propose to develop a cultured enterocyte model in which chylomicron assembly and transport can be studied in detail. Second, our data indicate the PITPalpha-deficient neurons exhibit intrinsic defects; most likely in signaling through the so-called 'survival pathways'. Using genetic, biochemical and signaling approaches we will test this hypothesis in the PITPalpha-/- model. These studies will be complemented by exploitation of a novel Drosophila system for study of PITPct signaling functions. Third, we will use genome engineering approaches to functionally dissect the physiological functions of individual PITPalpha phospholipid transfer activities. In particular, we will test whether the key physiological function of PITPalpha is to facilitate phosphoinositide synthesis and, if so, what specific phosphoinositides are regulated in a PITPalpha-dependent manner and what signaling pathways are involved. PITPs play central roles in regulating signal transduction pathways that interface with diverse cellular processes. We now establish that these functional interfaces are relevant to human chylomicron retention disease, disorders of glucose homeostasis, and spinocerebellar degeneration. The Bankaitis laboratory is uniquely poised to address questions of mechanism of PITPalpha function as it has developed unique experimental systems for analysis.