Gene-regulatory networks in podocytes in health and disease

• Applicants: Professor Dr. Andreas Beyer; Dr. Martin Kann
• Subject Area: Nephrology
• Term: from 2016 to 2019
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 315284411

Podocytes are key to maintaining the filtration function of our kidneys. Damage to podocytes results in urinary protein excretion, which in turn leads to progression of kidney disease. In response to damage, podocytes employ mechanisms involving the cell nuclei by changing their gene expression pattern and by activating pathways that signal to the nucleus. However, despite longstanding knowledge of hereditary podocyte disease caused by mutations in nuclear proteins such as WT1, such nuclear mechanisms in podocytes are not well understood. In this context, the transcription factor (TF) WT1 plays an important role as it is indispensable to podocyte development and maintenance. Thus, we have previously characterized the WT1-dependent podocyte gene regulatory network in mice in vivo. In this network, WT1 acts as a pioneering TF and is bound to the genes of further podocyte TF such as Mafb and Lmx1b. Furthermore, in a podocyte disease model, genes relevant to podocyte damage pathways are differentially bound by WT1. Based on these findings, we now aim to test the hypothesis that in healthy podocytes, WT1 orchestrates the binding of further podocyte TFs, which in turn assume specific functions in podocyte physiology. To this end, we will employ chromatin immunoprecipitation (ChIPseq) to characterize binding sites various TFs relevant to podocytes in genome wide fashion. Gene expression analysis of mouse podocytes by RNA sequencing (RNAseq) will complement this data and identify TF-specific gene regulatory effects on defined podocyte components. The integration of these datasets with the existing WT1 network by bioinformatic analyses will allow for detailed insight into the function of TFs in podocyte maintenance as well as pathogenetic mechanisms in hereditary podocyte disease. In a second approach, we aim to investigate how WT1 and the physiologic podocyte TF network interact with the nuclear effectors of pathways activated upon podocyte damage. To this end, we will use mouse models of genetic and toxic podocyte damage and assess effector and WT1 binding sites by ChIPseq at an early and late stage of podocyte disease. In parallel, we will assess podocyte gene expression changes with disease by RNAseq. Bioinformatic analyses of all datasets will the enable us to identify dependencies between physiologic and pathologic gene regulatory networks, and to characterize a disease stage specific podocyte response on a nuclear level in a systems biology approach. We anticipate gaining significant insight into reprogramming of gene regulatory networks upon podocyte damage by both WT1 as well as further TFs relevant to podocyte maintenance. Furthermore, we will identify the effects of such reprogramming focusing on podocyte signaling pathways with an established role in podocyte damage. These findings will therefore shed light on established and novel mechanisms in podocyte disease.

• DFG Programme: Research Grants