REGULATION AND SIGNALING OF PKD2 CATION CHANNEL IN SPERM

精子中 PKD2 阳离子通道的调控和信号传导

• 批准号: 7301078
• 负责人: XIANGYI LU
• 金额: $ 20.6万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-03-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7301078
• 关键词: Drosophilidae; autosomal dominant trait; biological models; biological signal transduction; calcium binding protein; calcium channel; calcium flux; cations; cilium /flagellum motility; electrospray ionization mass spectrometry; epithelium; gene mutation; immunocytochemistry; intermolecular interaction; phenotype; phosphoproteins; phosphorylation; polycystic kidney; protein kinase A; protein localization; protein quantitation /detection; protein structure function; proteomics; recombinant proteins; sperm; sperm motility

DESCRIPTION (provided by applicant): Autosomal dominant polycystic kidney disease (ADPKD) is characterized by kidney cyst formation that progresses to end-stage renal failure at a high frequency. The disease also encompasses systemic vascular complications such as hypertension and vascular aneurysms that increase the overall mortality of the disease. Mutations in two genes Pkd1 and Pkd2 account for most cases of ADPKD. Recent studies show that PKD2 functions as a calcium-activated, calcium-permeable cation channel; and PKD1 plays a role in regulating the PKD2 channel activity. A high concentration of PKD1 and PKD2 co-localizes on primary cilia of renal epithelia. The current working model is that cilium-associated PKD1/PKD2 protein complex mediates Ca2+ influx through cilia bending in response to fluid flow in the renal tubules. Loss of either PKD1 or PK02 leads to a common defect in ciliary Ca2+ influx and this is one of the contributing factors of cyst formation in the kidney. This proposal utilizes a novel ciliary model of PKD2 in Drosophila sperm to substantiate the current model of PKD2 as cilium-associated, mechano-/chemo-sensitive cation channel. We show that Drosophila PKD2 (CG6504) is a conserved cation channel of the PKD2 family that localizes to sperm flagellar cilia where it is required for responding to signals that regulate asymmetric flagellar motility and directional sperm movement. This Drosophila PKD2 function appears to be analogous to mammalian PKD2 function in the motile nodal cilia. Directional beating of nodal cilia, which contain PKD2, is required for left-right axis determination in mammals. The goals of this proposal are to elucidate PKD2 channel regulation and signaling through structure-function analyses of the Drosophila PKD2 protein, phosphoproteomic analyses of PKD2-dependent downstream protein phosphorylation. and genetic analyses of candidate kinases and phosphoproteins in the pathway. The proposed research will identify structural elements involved in PKD2 channel regulation and function in vivo in a ciliary context, and will also identify PKD2-dependent downstream phosphoproteins and use them to delineate a genetic pathway for PKD2 signaling and function.

描述（由申请人提供）：常染色体显性多囊肾病（ADPKD）的特征是肾囊肿形成，并以高频率进展为终末期肾衰竭。该疾病还包括全身血管并发症，例如高血压和血管动脉瘤，这些并发症增加了该疾病的总体死亡率。大多数 ADPKD 病例是由 Pkd1 和 Pkd2 两个基因突变引起的。最近的研究表明 PKD2 具有钙激活、钙渗透性阳离子通道的功能； PKD1对PKD2通道活性具有调节作用。高浓度的 PKD1 和 PKD2 共定位于肾上皮的初级纤毛上。目前的工作模型是纤毛相关的 PKD1/PKD2 蛋白复合物响应肾小管中的液体流动，通过纤毛弯曲介导 Ca2+ 流入。 PKD1 或 PK02 的缺失会导致睫状 Ca2+ 内流的常见缺陷，这是肾脏囊肿形成的促成因素之一。该提案利用果蝇精子中 PKD2 的新型纤毛模型来证实 PKD2 的当前模型作为纤毛相关的、机械/化学敏感的阳离子通道。我们发现，果蝇 PKD2 (CG6504) 是 PKD2 家族的保守阳离子通道，定位于精子鞭毛纤毛，在此处，它需要响应调节不对称鞭毛运动和定向精子运动的信号。果蝇 PKD2 的这种功能似乎与哺乳动物运动结纤毛中的 PKD2 功能类似。含有 PKD2 的结纤毛的定向跳动是确定哺乳动物左右轴所必需的。该提案的目标是通过果蝇 PKD2 蛋白的结构功能分析、PKD2 依赖性下游蛋白磷酸化的磷酸化蛋白质组学分析来阐明 PKD2 通道调节和信号传导。以及该途径中候选激酶和磷蛋白的遗传分析。拟议的研究将鉴定在睫状体环境中参与体内 PKD2 通道调节和功能的结构元件，还将鉴定 PKD2 依赖性下游磷蛋白，并利用它们来描绘 PKD2 信号传导和功能的遗传途径。