Project Summary A limited understanding of clinically relevant signaling pathways has limited the development of therapeutic agents for human glomerular disease. Our long-term goal is to identify novel therapeutic targets for the treatment of glomerular disease by elucidating the details and functional significance of key signaling pathways that regulate podocyte injury and survival. Our preliminary data have shown that YAP silencing disrupts podocyte focal adhesion architecture and actin cytoskeletal integrity. Using RNA-Seq to compare the transcriptomic profile of control and YAP knockdown mouse podocytes, the most significantly enriched gene ontology molecular function term among the differentially upregulated genes is “potassium channel regulator activity”. YAP silencing significantly increases KCNN4 gene and encoded KCa3.1 (intermediate-conductance calcium-activated potassium channel) protein expression. KCa3.1 inhibitors decrease podocyte actin cytoskeletal disruption, pathogenic intracellular calcium signaling and potassium efflux along with decreasing podocyte injury in an acute in vivo model of glomerular disease. The overall objective of this application is to define the role of YAP in podocyte survival and as a potential therapeutic target in proteinuric kidney disease. Our hypothesis is that YAP nuclear expression is essential for maintaining the integrity of the podocyte actin cytoskeleton, including through repression of KCNN4 gene expression. The loss or inactivation of YAP enhances pathogenic KCa3.1 signaling in podocytes. Our hypothesis will be tested by pursuing two specific aims: Aim 1 will define the mechanism of Hippo-YAP regulation of podocyte KCa3.1 function. We will define the mechanism by which Hippo-YAP regulates KCa3.1 channel function in podocytes and the mechanism of podocyte injury induced by calcium-activated potassium channel activity. We will also test the in vivo interaction and functional role of YAP inhibition of KCNN4/KCa3.1 function. In Aim 2, we will develop and test the efficacy of KCa3.1 inhibitors for podocyte protection. We will use rational design and medicinal chemistry to optimize KCa3.1 inhibitors for podocyte protection, test the ability of novel KCa3.1 inhibitors to protect podocytes from injury while decreasing pathogenic potassium efflux and intracellular calcium signaling, and test the ability of KCa3.1 inhibitors to decrease albuminuria and slow kidney disease progression in relevant in vivo glomerular disease models. Currently, there are no KCa3.1 inhibitors in clinical development for podocytopathies, FSGS or proteinuric kidney disease. The work proposed is expected to characterize the role of Hippo-YAP signaling in regulating KCa3.1 function in podocytes while developing a novel therapeutic strategy for proteinuric kidney disease.