Focal segmental glomerulosclerosis (FSGS) is a frequent and severe glomerular disease characterized by destabilization of podocyte foot processes. We report that transgenic expression of the microRNA miR-193a in mice rapidly induces FSGS with extensive podocyte foot process effacement. Mechanistically, miR-193a inhibits the expression of the Wilms' tumor protein (WT1), a transcription factor and master regulator of podocyte differentiation and homeostasis. Decreased expression levels of WT1 lead to downregulation of its target genes PODXL (podocalyxin) and NPHS1 (nephrin), as well as several other genes crucial for the architecture of podocytes, initiating a catastrophic collapse of the entire podocyte-stabilizing system. We found upregulation of miR-193a in isolated glomeruli from individuals with FSGS compared to normal kidneys or individuals with other glomerular diseases. Thus, upregulation of miR-193a provides a new pathogenic mechanism for FSGS and is a potential therapeutic target.

The Wilms' tumor suppressor gene 1 (WT1) encodes a zinc finger transcription factor. Mutation of WT1 in humans leads to Wilms' tumor, a pediatric kidney tumor, or other kidney diseases, such as Denys-Drash and Frasier syndromes. We showed previously that inactivation of WT1 in podocytes of adult mice results in proteinuria, foot process effacement, and glomerulosclerosis. However, the WT1-dependent transcriptional network regulating podocyte development and maintenance in vivo remains unknown. Here, we performed chromatin immunoprecipitation followed by high-throughput sequencing with glomeruli from wild-type mice. Additionally, we performed a cDNA microarray screen on an inducible podocyte-specific WT1 knockout mouse model. By integration of cistromic and transcriptomic analyses, we identified the WT1 targetome in mature podocytes. To further analyze the function and targets of WT1 in podocyte maturation, we used an Nphs2-Cre model, in which WT1 is deleted during podocyte differentiation. These mice display anuria and kidney hemorrhage and die within 24 hours after birth. To address the evolutionary conservation of WT1 targets, we performed functional assays using zebrafish as a model and identified Nphs2, Mafb, and Magi2 as novel WT1 target genes required for podocyte development. Our data also show that both Mafb and Magi2 are required for normal development of the embryonic zebrafish kidney. Collectively, our work provides insights into the transcriptional networks controlled by WT1 and identifies novel WT1 target genes that mediate the function of WT1 in podocyte differentiation and maintenance.

The Wilms' tumor protein Wt1 plays an essential role in mammalian urogenital development. WT1 mutations in humans lead to a variety of disorders, including Wilms' tumor, a pediatric kidney cancer, as well as Frasier and Denys-Drash syndromes. Phenotypic anomalies in Denys-Drash syndrome include pseudohermaphroditism and sex reversal in extreme cases. We have used cDNA microarray analyses on Wt1 knockout mice to identify Wt1-dependent genes involved in sexual development. The gene most dramatically affected by Wt1 inactivation was Amhr2, encoding the anti-Müllerian hormone (Amh) receptor 2. Amhr2 is an essential factor for the regression of the Müllerian duct in males, and mutations in AMHR2 lead to the persistent Müllerian duct syndrome, a rare form of male pseudohermaphroditism. Here we show that Wt1 and Amhr2 are coexpressed during urogenital development and that the Wt1 protein binds to the promoter region of the Amhr2 gene. Inactivation and overexpression of Wt1 in cell lines was followed by immediate changes of Amhr2 expression. The identification of Amhr2 as a Wt1 target provides new insights into the role of Wt1 in sexual differentiation and indicates, in addition to its function in early gonad development and sex determination, a novel function for Wt1, namely, in Müllerian duct regression.

The Wilms tumor protein WT1 is involved in the development of several organs, including the gonads. WT1 mutations in humans lead to syndromes associated with impaired sexual development and Wt1 knockout mice show regression of gonad anlagen. As a transcription factor, WT1 fulfills its function by regulating a set of target genes. With respect to gonad development only few in vivo WT1 targets, e.g. steroidogenic factor 1 (SF1) have been identified so far. To get a comprehensive view of WT1 targets in the gonad, we compared gene expression in urogenital ridges of wild-type and Wt1^–/– embryos. We found almost 150 genes differentially expressed higher than factor three, using microarray analysis. To confirm these results we performed quantitative real-time RT-PCR for many genes and observed a high degree of concordance between microarray and real-time RT-PCR results. Employing in situ hybridization we found ‘WT1 activated genes’ to be expressed in gonads, mesonephroi and coelomic epithelium – those parts of the urogenital ridge with Wt1 expression. Interestingly, many of the differentially expressed genes are known to show sex-specific expression at later time-points. These results provide a basis for investigation of developmental pathways in the urogenital ridge downstream of WT1 and for identification of new candidate genes involved in early urogenital ridge development. For example we provide a first potential target of WT1 in the coelomic epithelium – Muc16, and a gene regulated by the WT1 target SF1 – Gata4.

Hedgehog (Hh) signaling has been implicated in the development of osteoblasts and osteoclasts whose balanced activities are critical for proper bone formation. Since many mouse mutants