Internal Medicine D, and.
Institutes of Cell Dynamics and Imaging.
J Am Soc Nephrol. 2019 Jun;30(6):1006-1019. doi: 10.1681/ASN.2018040362. Epub 2019 May 16.
Patients with certain mutations in the gene encoding the slit diaphragm protein Nephrin fail to develop functional slit diaphragms and display severe proteinuria. Many adult-onset glomerulopathies also feature alterations in Nephrin expression and function. Nephrin signals from the podocyte slit diaphragm to the Actin cytoskeleton by recruiting proteins that can interact with C3G, a guanine nucleotide exchange factor of the small GTPase Rap1. Because Rap activity affects formation of focal adhesions, we hypothesized that Nephrin transmits signals to the Integrin receptor complex, which mediates podocyte adhesion to the extracellular matrix.
To investigate Nephrin's role in transmitting signals to the Integrin receptor complex, we conducted genetic studies in Drosophila nephrocytes and validated findings from Drosophila in a cultured human podocyte model.
Drosophila nephrocytes form a slit diaphragm-like filtration barrier and express the Nephrin ortholog Sticks and stones (Sns). A genetic screen identified as necessary for nephrocyte function. , nephrocyte-specific gene silencing of or compromised nephrocyte filtration and caused nephrocyte diaphragm defects. Nephrocytes with impaired Sns or C3G expression displayed an altered localization of Integrin and the Integrin-associated protein Talin. Furthermore, gene silencing of partly rescued nephrocyte diaphragm defects of an overexpression phenotype, pointing to genetic interaction of and in nephrocytes. We also found that activated Nephrin recruited phosphorylated C3G and resulted in activation of Integrin 1 in cultured podocytes.
Our findings suggest that Nephrin can mediate a signaling pathway that results in activation of Integrin 1 at focal adhesions, which may affect podocyte attachment to the extracellular matrix.
编码裂孔隔膜蛋白 Nephrin 的基因突变的患者无法形成功能性裂孔隔膜，并表现出严重的蛋白尿。许多成人发病的肾小球病变也表现出 Nephrin 表达和功能的改变。Nephrin 通过募集可以与小 GTPase Rap1 的鸟嘌呤核苷酸交换因子 C3G 相互作用的蛋白，从足细胞裂孔隔膜向肌动蛋白细胞骨架传递信号。由于 Rap 活性影响粘着斑的形成，我们假设 Nephrin 将信号传递到整合素受体复合物，该复合物介导足细胞与细胞外基质的黏附。
为了研究 Nephrin 在向整合素受体复合物传递信号中的作用，我们在果蝇肾细胞中进行了遗传研究，并在培养的人足细胞模型中验证了果蝇的研究结果。
果蝇肾细胞形成类似于裂孔隔膜的滤过屏障，并表达 Nephrin 同源物 Sticks and stones (Sns)。遗传筛选确定 是肾细胞功能所必需的。特异性基因沉默 或 会损害肾细胞的滤过功能，并导致肾细胞隔膜缺陷。表达受损的 Sns 或 C3G 的肾细胞显示出整合素和整合素相关蛋白 Talin 的定位改变。此外，基因沉默 部分挽救了 过表达表型的肾细胞隔膜缺陷，表明 与 之间在肾细胞中有遗传相互作用。我们还发现，激活的 Nephrin 募集磷酸化的 C3G，导致培养的足细胞中整合素 1 的激活。
我们的研究结果表明，Nephrin 可以介导一种信号通路，导致整合素 1 在粘着斑处激活，这可能影响足细胞与细胞外基质的附着。
