Novel PC2 regulation of ezrin in renal epithelia reveals insights into ADPKD cystogenesis

Abstract

Autosomal dominant polycystic kidney disease is caused by the loss of function of either two transmembrane proteins, polycystin-1 or polycystin-2. In renal epithelia, the consequence of polycystin loss is the formation of progressive, focal, fluid-filled cysts. However, the function and associated downstream signaling pathways specific to the polycystins have not been defined. Therefore, a new in vitro tubuloid model was designed to investigate the proximate cellular changes in renal epithelial cells following inactivation of Pkd2, the gene that encodes for polycystin-2. This model system reinforced the relevance of proteins associated with cell junctions, adhesions, and matrix in the cyst mechanism. The impact of this model was further supported through morphometrical analysis of epithelial compartmentalization in human ADPKD tissue, demonstrating an altered apical compartment in emerging cysts compared to noncystic tubules. Seeking connection between the junctions and disrupted apical compartment led to investigation of ezrin, a master scaffold in the apical compartment in renal epithelial cells. Ezrin plays a critical role in regulation of polarity, cytoskeleton organization, and protein trafficking, and the downstream consequences of its disruption have not been elucidated. Investigation into the initiating events of cystogenesis in ADPKD revealed a dramatic change in ezrin, following loss of PC2 in our tubuloid model, cystic mouse model, and pathological human ADPKD tissue. Based on this novel regulatory relationship between PC2 and ezrin, as well as the antecedent loss of ezrin to cyst formation in mice, ezrin was overexpressed in the pkd2 morpholino zebrafish model. Increased expression of ezrin diminished the formation of pronephric cysts. This lead to the design of a cyst rescue mouse model, which has exhibited promising preliminary data for cyst area reduction with additional ezrin. The disruption of ezrin in Pkd2 inducible in vitro and in vivo model systems, changes in ADPKD patient tissue, and rescue of pronephric cysts in the pkd2 MO suggest there is a role of ezrin in renal cystogenesis. Understanding the relationship of ezrin, with PC2 in renal epithelial cells will help elucidate the mechanism of ADPKD cystogenesis and define important downstream pathways necessary for epithelial functions.