Establishing an in vivo functional analysis system for renal gene discovery in Drosophila pericardial nephrocytes. Fujian Zhang¹, Ying Zhao¹, Zhe Han^1,2. 1) Department of Internal Medicine, Division of Molecular Medicine and Genetics, University of Michigan, Ann Arbor, MI; 2) Department of Cell and Developmental Biology, University of Michigan, Ann Arbor, MI.

   The glomerular podocyte plays a central role in the mammalian renal system. Most known renal disease genes are involved in podocytes function, but understanding of the podocyte biology has been hindered by the low accessibility of mammalian nephrons in vivo. The Drosophila nephrocyte shares remarkable similarities to the glomerular podocyte, making it a potential ideal model to study podocyte biology. However, the lack of functional readout for nephrocytes makes it hard to exploit the power of Drosophila genetics. Here, we present a novel functional analysis of nephrocytes and established an in vivo genetic screen system for renal gene discovery. We found that nephrocytes efficiently uptake secreted fluorescent protein. We generated a transgenic line carrying secreted fluorescent protein that is accumulated in nephrocytes, and combined it to a nephrocyte specific driver for targeted gene knockdown to identify genes required for nephrocyte function. To validate this system, we examined the effects of knocking down sns and duf, the Drosophila homologues of nephrin and Neph1, respectively, in pericardial nephrocytes. We found that sns or duf knockdown completely abolished ANF-RFP protein accumulation in pericardial nephrocytes. Ultra-structure analysis demonstrated that sns is required for nephrocyte diaphragm and lacunar structure formation that are essential for protein uptake. Our preliminary genetic screen also identified Mec2, which encodes the homologue of mammalian Podocin, another slit diaphragm component linked to renal disease. These findings suggested that the functional analysis system we developed has made the Drosophila pericardial nephrocyte an ideal in vivo model to help identify genes involved in podocyte biology and to facilitate the renal disease gene discovery.