Kidney development and cystogenesis in Medaka

描述(申请人提供)：常染色体显性遗传性多囊肾病(ADPKD)是一种遗传性疾病，在人类中发生的频率为1：1000，其特征是肾小管上囊肿形成，液体运输失控和细胞外黏附改变。ADPKD是由分别编码多囊蛋白-1和多囊蛋白-2的PKD1或PKD2基因突变引起的。ADPKD囊性形成的主要原因和机制仍不清楚。我们试图在斑马鱼中敲除pkdl的尝试失败了，因为基因冗余。青竹是一种独特的近亲繁殖水生脊椎鱼模型系统，与斑马鱼相比，它在基因数量上表现出较低的冗余度。青竹的使用为开发研究器官发生和了解囊变机制的新系统提供了机会。鉴于PKD1突变占人类ADPKD病例的85%，我们以青竹中的pkd1为靶点来了解其功能(S)。初步结果表明，靶向青竹的pkdl(与斑马鱼相反)会导致前肾囊的形成。我们还建立了一个利用青竹研究肾脏、膀胱发生的系统，该系统对多囊蛋白-1相互作用蛋白的缺陷很敏感，如多囊蛋白-2，可用于评估大量多囊蛋白-1相互作用蛋白在体内的相关性。 在特定目标1中，我们将通过破坏pkdl mRNA剪接来测试特定多囊蛋白-1基序的体内功能，并在青竹中产生一系列等位基因缺失。为此，我们建议使用我们的泌尿生殖器官特异的青竹转基因GFP系来监测包囊形成的进展，并分析肾脏的表型以及由延迟引起的其他潜在的器官发生缺陷。在特定的目标2中，我们将验证多囊蛋白-1相互作用蛋白的生物学意义。

DESCRIPTION (provided by applicant): Autosomal dominant polycystic kidney disease (ADPKD) is a hereditary disease occurring at a frequency of 1:1000 in humans and characterized by cyst formation in kidney tubules, deregulated fluid transport and alteration of extracellular adhesion. ADPKD is caused by mutations in the PKD1 or PKD2 gene that encode polycystin-1 and polycystin-2, respectively. The primary causes and mechanisms of cyst formation in ADPKD remain elusive. Our attempts to knockdown pkdl in zebrafish failed because of gene redundancy. Medaka is a unique inbred aquatic vertebrate fish model system, which shows lower redundancy in gene number compared to zebrafish. The use of medaka provides an opportunity to develop a novel system to study organogenesis and to understand the mechanisms of cystogenesis. Given that PKD1 mutations account for 85% of all ADPKD cases in humans, we have targeted pkdl in medaka to understand its function(s). Preliminary results demonstrate that targeting pkdl in medaka (in contrast to zebrafish) causes prronephric cysts formation. We have also established a system using medaka for studies of kidney, cystogenesis that is sensitive for the defect of polycystin-1 interacting proteins such as polycystin-2, which can be used to assess the in vivo relevance of the large number of polycystin-1 interacting proteins. In specific aim 1, we will test the in vivo function of specific polycystin-1 motifs by disrupting pkdl mRNA splicing and generate an allelic series of deletions in medaka. For this purpose, we propose to use our urogenital specific medaka transgenic GFP line to monitor the progress of cyst formation, and to analyze the kidney phenotypes as well as other potential defects in organogenesis caused by the defers. In specific aim 2, we will validate the biological significance of polycystin-1 interacting proteins.