Impact of phosphatidylinositol metabolism for ciliary biology and renal function

慢性肾脏病是一种常见病，其发病率在全球范围内呈上升趋势。与其病因无关，肾功能受损的进展是基于纤维化的重塑过程。然而，潜在的分子因素和机制却知之甚少。单基因孟德尔病伴肾脏受累提供了识别这些未知因素的独特可能性，并提供了进一步了解此发病机制的分子机制。初级纤毛是以毛状细胞微管为基础的细胞器，存在于人体几乎所有静止细胞的表面，对细胞的各种基本功能具有重要作用。纤毛蛋白编码基因的突变会导致一系列疾病，这些疾病统称为“纤毛疾病”。表型谱以早期肾间质纤维化和肾小管基底膜崩解为特征。肾病相关纤毛病(NPHP-RC)一词概括了一组罕见的常染色体隐性遗传性肾脏疾病，这些疾病在生命的前三十年中占终末期肾脏疾病的大部分。通过对一例NPHP-RC患者的外显子全基因组测序，我们发现了导致睫毛功能障碍的PIK3C2A纯合子截短突变。该酶属于磷脂酰肌醇-3-激酶家族，在细胞生理中起着至关重要的作用，如内吞和胞吐。PIK3C2A位于纤毛基底部附近，其底物定位于纤毛轴丝内。然而，它的睫毛功能仍不清楚。本项目计划研究PIK3C2a及其脂类产物的细胞和纤毛功能，以及在CKD和纤毛疾病的病理生理学中的重要性。因此，我们建立了纯合子、杂合子和野生型等位基因携带者以及不同分子原因的NPHP-RC患者的原代细胞培养。这一资源使我们能够研究激酶在纤毛运输过程和纤毛信号转导中的作用。该酶及其产物的肾脏和肾外功能将通过使用低形态的Pik3c2a小鼠模型在体内进行研究。为了检查肌醇磷脂途径的一般肾脏和睫毛功能，该项目将得到进一步的小鼠模型(Inpp5e)的补充。这个项目将扩大对纤毛生物学和肾脏重塑过程的理解，这些过程可能与肾脏疾病有关。

Chronic kidney disease is a common disease and its prevalence is increasing worldwide. Independently of its etiology the progression of impaired kidney function is based on fibrotic remodeling processes. However, the underlying molecular factors and mechanisms are poorly understood. Monogenic, mendelian disorders with renal involvement offer the unique potential to identify these unknown factors and to provide further understanding of the molecular mechanisms involved in this pathogenesis. Primary cilia are hairlike cellular microtubule based organelles that exist on the surface of almost every quiescent cell of the human body and owe substantial importance for diverse essential cellular functions. Mutations in genes that encode for ciliary proteins lead to a broad complex of disorders, which have collectively been termed “ciliopathies”. The phenotypic spectrum is characterized by early renal interstitial fibrosis and tubular basement membrane disintegration. The term nephronophthisis-related ciliopathies (NPHP-RC) summarizes a group of rare autosomal-recessive kidney diseases which account for the majority of genetically caused end-stage renal disease during the first three decades of life. By whole exome sequencing of one affected individual with NPHP-RC we identified a homozygous truncating mutation in PIK3C2A leading to ciliary dysfunction. The enzyme belongs to the family of phosphatidylinositol - 3-kinases, which serve crucial roles in cell physiology, such as endo- and exocytosis. PIK3C2A was detected in close proximity to the ciliary base and its substrate localizes within the ciliary axoneme. However, its ciliary function remains unknown. This project plans to investigate the cellular und ciliary function of PIK3C2A and its lipid products as well as the importance for the pathophysiology of CKD and ciliopathies. Therefore, we established primary human cell cultures of the homozygous, the heterozygous and wildtype allele carrier as well as of affected individuals with NPHP-RC of different molecular causes. This resource enables us to examine the role of the kinase for periciliary transport processes and ciliary signaling. The renal and extra-renal function of the enzyme and its products will be investigated in vivo by using a hypomorphic Pik3c2a mouse model. In order to examine the general renal und ciliary function of the phosphoinositide pathway the project will be complemented by a further mouse model (Inpp5e). This project will extend the understanding of ciliary biology and renal remodeling processes which may have relevance to kidney dieases in general.