Molecular modulators of polycystin signaling

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Autosomal dominant polycystic kidney disease (ADPKD) is a common genetic disease that results from mutations in either of two proteins, polcysytin-1 (PC1) or polycystin-2 (PC2). More than two decades have passed since the genes encoding these proteins were discovered and there has been significant progress in understanding the functions of polycystins and their associated disease. Nonetheless, there remain substantial gaps in knowledge and lack of consensus about the precise functions of PC1 or PC2 and the mechanisms of ADPKD. Resolution of these gaps is of great significance given our expectation that optimal therapies for ADPKD are best developed based on the fundamental understanding of polycystin function in the mammalian kidney. Much of the current mechanistic understanding of polycystin function is based on studies of candidate pathways drawn from amongst known cellular mechanisms associated with functions such as differentiation, proliferation, transport and signaling. The lack of coalescence toward an interrelated unifying functional pathway in polycystin biology and the persistence of gaps in understanding of in vivo polycystin function despite extensive investigation suggests that the critical components of the most proximal polycystin signaling cascade have yet to be identified. Indeed, the polycystins were discovered as complex, entirely novel proteins and it stands to reason that they may function in a signaling pathway that is not among those that are currently well understood or studied. We made use of this concept in by applying an unbiased in vivo transcriptomic study using Translating Ribosome Affinity Purification (TRAP) RNASeq. From this, we identified upregulation of cell cycle and down regulation of oxidative phosphorylation as key pathways alterations in vivo. Among these, we found genotype dependent upregulation of a cilia associated transcription factor, Glis2, not previously considered to function in polycystin signaling or ADPKD pathogenesis. We made double mutants of Pkd1 with Glis2 in early onset and adult models and found Glis2 dosage-dependent rescue of cyst formation in both. Based on these findings we propose that Glis2 is a candidate for a downstream effector of PC1 function that is critical for cyst progression in ADPKD. In this study, we will determine the in vivo mechanism of action of Glis2 and establish its effects on cyst cell proliferation, apoptosis and ADPKD due to Pkd2. We will determine whether Glis2 is a target for therapy through both genetic and pharmacotherapeutic studies. We will assess whether in vivo genotype dependent transcriptional changes we have identified are similarly extended to cell culture systems with Pkd mutant genotypes. We will also evaluate the functional properties of Glis2 protein in Pkd mutant cell lines. In aggregate, these studies open a new direction of investigation for polycystin signaling and ADPKD pathogenesis.