Linking polarity signaling to mechanotransduction in glomerular podocytes

肾脏滤过屏障疾病是慢性肾脏疾病的主要原因，影响全球超过5亿人。不管根本原因是什么，肾小球损伤的程度总是由肾小球上皮细胞（足细胞）的损伤来确定的。后者是肾小球高度特化的上皮细胞，它们与有孔内皮细胞和肾小球基底膜一起构成肾脏滤过屏障。足细胞表现出独特的形态，形成初级和次级足突，完全包裹肾小球毛细血管。损伤后，这种特化的功能性形态发生改变，足细胞类似于经典的柱状上皮细胞。其直接后果是足细胞的细胞形状改变，细胞丢失到原发尿液中。鉴于足细胞有丝分裂后的性质，任何细胞的损失都不能通过邻近细胞的增殖来弥补，使毛细血管空白，并导致大量蛋白质损失进入初级尿液。肌动蛋白-细胞骨架在这一去分化过程中发挥着重要作用，但其调控途径要么尚未被确定，要么尚未充分表征。损伤小鼠原代足细胞的MS/MS初步数据显示，损伤后推测的机械传感器蛋白丝蛋白B显著增加。在极性信号机制缺陷的果蝇肾细胞中，其同源物Cher也显著增加，这表明足细胞中机械转导过程和细胞极性信号通路不仅非常重要，而且这两种途径都可能存在遗传依赖性。因此，在本项目提案中，我们的目标是研究机械转导和细胞极性信号传导对足细胞生物学的影响。我们将利用果蝇肾细胞模型进一步揭示机械传感器蛋白丝蛋白的功能作用。为此，我们将产生不同的转基因果蝇，并表征不同丝蛋白结构域对肾细胞形态和功能的贡献。在第二种独立的方法中，我们将研究异常细胞极性信号传导对果蝇肾细胞体内机械转导过程的影响。为了进一步研究这两种通路之间的联系，我们还将分析受干扰的机械感觉对肾细胞细胞极性的影响。在这项提议中，我们不仅旨在揭示丝蛋白在体内的功能作用，而且旨在表征体内肾细胞中机械转导和细胞极性信号传导之间的潜在联系。本研究结果将有助于了解足细胞损伤后形态变化的潜在调控机制。基于这些已获得的知识，未来可以开发治疗肾小球疾病的新疗法。

Diseases of the kidney filtration barrier are a major cause of chronic kidney disease affecting more than 500 million people worldwide. Independent of the underlying cause, the degree of glomerular injury is always defined by the damage to the epithelial cells of the glomerulus, the podocytes. The latter are highly specialized epithelial cells of the glomerulus, which build the actual kidney filtration barrier together with fenestrated endothelial cells and the glomerular basement membrane. Podocytes exhibit a unique morphology and form primary and secondary foot processes, which enwrap the glomerular capillaries completely. Upon injury this specialized and functional morphology alters and podocytes resemble classical columnar epithelial cells. As a direct consequence, the podocytes’ cell shape changes and the cells are lost into the primary urine. Given the post mitotic nature of podocytes any cellular loss cannot be compensate by proliferation of neighboring cells leaving the capillaries blank and causing immense loss of protein into the primary urine. The actin-cytoskeleton plays an important role during this dedifferentiation process, but the regulatory pathways are either not identified so far or only characterized insufficiently. Preliminary MS/MS data of injured primary mouse podocytes revealed a significant increase of the putative mechanosensor protein Filamin B upon injury. Its Drosophila homolog Cher was also significantly increased in Drosophila nephrocytes with a defective polarity signaling machinery, suggesting not only a high importance of mechanotransduction processes and cell polarity signaling pathways in podocytes, but also a potential genetic dependency of both pathways. Therefore, within this project proposal we aim to investigate the impact of mechanotransduction and cell polarity signaling on podocyte biology. We will make use of the Drosophila nephrocyte model to further unravel the functional role of the mechanosensor protein Filamin. To this end, we will generate different transgenic flies and characterize the contribution of the different Filamin domains for nephrocyte morphology and function. In a second and independent approach, we will study the impact of aberrant cell polarity signaling on mechanotransduction process in Drosophila nephrocytes in vivo. To further examine the interconnection between the two pathways, we will also analyze effects of disturbed mechanosensation on cell polarity in nephrocytes. Within this proposal, we not only aim to unravel the functional role of Filamin in vivo, but also aim to characterize the potential interconnection between mechanotransduction and cell polarity signaling in nephrocytes in vivo. The findings of this proposal will clearly help to understand the underlying mechanisms regulating morphological changes of podocytes upon injury. Based on this acquired knowledge novel therapies treating glomerular diseases can be developed in the future.