Role of PACS proteins in polycystin-2 trafficking and ADPKD

描述（由申请人提供）：该项目的长期目标是确定分选蛋白PACS-1和PACS-2在常染色体显性多囊肾病（ADPKD）中的作用。这种遗传性疾病表现为肾脏中形成大量囊肿，最终导致过度细胞凋亡和正常组织的破坏。 ADPKD 通常由多囊蛋白-2 突变引起，多囊蛋白-2 是一种在多个亚细胞器中发挥作用的钙渗透性离子通道。我们最近确定了分子运输机制 - PACS-1 和 PACS-2 - 控制着多囊蛋白-2 在内质网 (ER)、高尔基体和细胞表面之间的逐步运动。此外，我们发现PACS-1和PACS-2将蛋白质运输与细胞凋亡和细胞分化结合起来。我们假设 PACS-1 和 PACS-2 是多功能分选蛋白，控制正常肾脏中多囊蛋白-2 的亚细胞定位，并且 PACS-1 和 PACS-2 的错误调节导致多囊肾中观察到的囊肿发生和过度凋亡。我们确定 PACS-2 是第一个 COPI 连接器，目标 1 中的实验将确定 PACS-2 和 COPI 如何结合，将多囊蛋白-2 定位到 ER（该离子通道的主要细胞储存库）。此外，我们还表明 PACS-1 是一种 AP-1 连接器，可将多囊蛋白-2 定位到跨高尔基体网络 (TGN)。目标 2 中的研究将确定 PACS-1 和 PACS-2 分选活性的调节如何影响多个亚细胞区室中的多囊蛋白-2 钙峰值。尽管多囊蛋白-2 突变与 ADPKD 之间存在因果关系，但从通道功能障碍到囊肿发生和疾病的具体步骤仍知之甚少。我们最近发现 ADPKD 肾脏中的 PACS-1 表达严重降低，而 PACS-2 表达变化相对较小——这一组合有利于培养细胞的凋亡。目标 3 的研究将确定 ADPKD 肾脏中 PACS-1 和 PACS-2 的细胞表达，并将使用多囊肾病小鼠模型测试 PACS-2 的缺失是否会抑制囊肿发生。我们提出的研究的成功完成将首次阐明多功能运输机制——PACS-1和PACS-2——调节多囊蛋白-2在多个细胞器中传导钙电流的能力，以及PACS-1和PACS-2表达的错误调节如何导致ADPKD中发现的囊肿发生和过度凋亡。

DESCRIPTION (provided by applicant): The long-term goal of this project is to determine the role of the sorting proteins PACS-1 and PACS-2 in autosomal dominant polycystic kidney disease (ADPKD). This inherited disorder manifests in formation of numerous cysts in the kidney, culminating in excessive apoptosis and destruction of normal tissue. ADPKD is frequently caused by mutation of polycystin-2, a calcium-permeable ion channel that functions in multiple subcellular organelles. We recently identified the molecular trafficking machinery-PACS-1 and PACS-2- governing the stepwise movement of polycystin-2 between the endoplasmic reticulum (ER), Golgi and the cell surface. In addition, we discovered that PACS-1 and PACS-2 integrate protein trafficking with apoptosis and cell differentiation. We hypothesize that PACS-1 and PACS-2 are multifunctional sorting proteins that control the subcellular localization of polycystin-2 in the normal kidney, and that misregulation of PACS-1 and PACS-2 contributes to the cystogenesis and excess apoptosis observed in the polycystic kidney. We identified PACS-2 as the first COPI connector, and experiments in Aim 1 will determine how PACS-2 and COPI combine to localize polycystin-2 to the ER-the principle cellular reservoir for this ion channel. Also, we showed that PACS-1 is an AP-1 connector that localizes polycystin-2 to the trans-Golgi network (TGN). Studies in Aim 2 will determine how regulation of PACS-1 and PACS-2 sorting activity effects polycystin-2 calcium spikes in multiple subcellular compartments. Despite the causal relationship between polycystin-2 mutations and ADPKD, the steps leading from channel dysfunction to cystogenesis and disease are poorly understood. We recently found that PACS-1 expression is severely reduced in the ADPKD kidney, while PACS-2 expression is relatively little changed-a combination that favors apoptosis in cultured cells. Studies in Aim 3 will determine the cellular expression of PACS-1 and PACS-2 in the ADPKD kidney and will test whether loss of PACS-2 inhibits cystogenesis using a mouse model of polycystic kidney disease. Successful completion of our proposed studies will illuminate for the first time the multifunctional trafficking machinery- PACS-1 and PACS-2-that regulates the ability of polycystin-2 to conduct calcium currents in multiple organelles, and how misregulation of PACS-1 and PACS-2 expression contributes to the cystogenesis and excess apoptosis found in ADPKD.