Understanding Polycystin-1 Structure-GPCR Function for the Development of New Therapeutic Approaches in ADPKD

总结 常染色体显性遗传性多囊肾病（ADPKD）是最常见的潜在致死性遗传病， 疾病85%的ADPKD病例是由于PKD 1基因突变， 大约1/4 - 1/3是由于错义突变和小的框内缺失， 功能性救援PKD 1编码多囊蛋白-1（PC 1），这是一种大型复杂蛋白质，被认为是 非典型G蛋白偶联受体（GPCR）。现在已知PC 1-G蛋白调节的丧失是 PKD发病机制的基础，但我们基本上不知道PC 1的机制， 调节G蛋白信号传导活性。我们的长期目标是了解PC 1-G的潜在机制 蛋白调节，以便靶向和功能性地恢复或增强该功能作为ADPKD的治疗。 基于PC 1和GPCR的粘附类之间的结构和功能相似性， 从我们的初步研究有趣的结果，我们提出了一个机制，涉及一个神秘的拴系 肽配体负责通过PC 1调节G蛋白信号传导，并且还参与 肾囊肿的病因有哪些因此，本提案的主要目标是确定 PC 1介导的G蛋白信号转导的结构-功能关系 并证明这种调节机制在发病机制和潜在治疗中的相关性， ADPKD。具体来说，我们打算确定的拴系肽配体的PC 1的基本性质 负责G蛋白信号的调节，绘制参与拴系配体的PC 1区域， 依赖性信号传导，并确定肾的肽配体依赖性拯救的条件。 囊肿形成所提出的研究将采用已建立的细胞系沿着用标准的 分子生物学方法和生物化学测定来评估修饰的表达的信号传导能力 PC 1和可溶性栓系配体衍生肽的构建体，并将包括治疗囊性 在离体和体内模型系统中，用可溶性栓系配体衍生的肽对肾脏进行了研究。成功 该项目的完成有望揭示PC 1调控G蛋白信号传导的分子细节 通过其隐蔽的拴系配体，并证明这种调节机制作为一种新的 治疗靶点阐明PC 1调节G蛋白信号转导的机制将有助于我们进一步研究G蛋白信号转导的机制。 了解ADPKD的分子发病机制，这对于开发新的 用于治疗或预防这种疾病的疗法。

Summary Autosomal dominant polycystic kidney disease (ADPKD) is the most common potentially lethal genetic disease. Eighty-five percent of the cases of ADPKD are due to mutation of the PKD1 gene of which approximately ¼-1/3 are due to missense mutations and small, in-frame deletions that may be amenable to functional rescue. PKD1 encodes polycystin-1 (PC1), a large and complex protein thought to function as an atypical G protein-coupled receptor (GPCR). The loss of PC1-G protein regulation is now known to be fundamental to the pathogenesis of PKD, yet we know essentially nothing about the mechanism whereby PC1 regulates G protein signaling activity. Our long-term goal is to understand the mechanism underlying PC1-G protein regulation in order to target and functionally restore or augment this function as a treatment for ADPKD. Based on the structural and functional similarities between PC1 and the adhesion class of GPCRs, and intriguing results from our preliminary studies, we propose that a mechanism involving a cryptic tethered peptide ligand is responsible for the regulation of G protein signaling by PC1, and furthermore, is involved in the pathogenesis of renal cyst formation. The major goals of this proposal, therefore, are to determine the structure-function relationships behind the tethered ligand-mediated regulation of G protein signaling by PC1 and to demonstrate the relevance of this regulatory mechanism in the pathogenesis and potential treatment of ADPKD. Specifically, we intend to determine the essential properties of the tethered peptide ligand of PC1 responsible for its regulation of G protein signaling, map the regions of PC1 involved in tethered ligand- dependent signaling, and determine the conditions for the peptide ligand-dependent rescue of renal cystogenesis. The proposed studies will employ transfection of established cell lines along with standard molecular biology methods and biochemical assays to assess the signaling capability of modified expression constructs of PC1 and of soluble tethered ligand-derived peptides, and will include the treatment of cystic kidneys with soluble tethered ligand-derived peptides in both ex vivo and in vivo model systems. Successful completion of this project is expected to reveal molecular details of the regulation of G protein signaling by PC1 via its cryptic, tethered ligand, and to demonstrate the potential of this regulatory mechanism as a novel therapeutic target. Elucidating the mechanism of PC1-regulated G protein signaling will advance our understanding of the molecular pathogenesis of ADPKD, which is imperative for the development new therapies for the treatment or prevention of this disease.