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Cytokinesis, the final step of cell division, is a sequential event that results in the physical separation of two daughter cells. The process involves the following sequential steps a) specification of the division site, b) actomyosin ring assembly, c) actomyosin ring constriction, and d) cell separation. I used Fission yeast (Schizosaccharomyces pombe), a rod-shaped unicellular eukaryote, as the model organism to understand cytokinesis. The core cytokinetic machinery of this genetically tractable model organism is conserved in animal cells and higher eukaryotes. Cytokinesis failure is often associated with cancer, blood disorders, and developmental defects due to the accumulation of polyploid cells. Thus, it is essential to study the signaling pathways that regulate cytokinesis to improve human health and cure diseases.Calcium is an essential secondary messenger that plays a key regulatory role in various cellular processes, including cytokinesis. However, its functional role during cytokinesis remains underappreciated. Previous studies have demonstrated a transient localized increase of intracellular calcium (calcium spikes) during the cytokinesis of animal and fish embryos. However, such calcium transients only remained correlated to cytokinesis and the causal link was never established. Here I employed live-cell calcium imaging and quantitative image analysis to demonstrate the conserved nature of such calcium spikes during fission yeast cytokinesis. I discovered two calcium spikes, one at the start of cleavage furrow ingression and another after the daughter cell separation. I found that depletion of external calcium vastly reduced the cytokinetic calcium spikes but did not block them completely. The inhibition of the calcium spikes slowed the cleavage furrow ingression and led to frequent lysis of separating daughter cells, suggesting the importance of calcium spikes in maintaining the pace of actomyosin ring constriction and integrity of the separating daughter cells.In search of the potential mechanisms of genes controlling such cytokinetic calcium spikes, I examined the contribution from a putative calcium permeable TRP channel, Pkd2. I found that the inactivation of Pkd2 reduced intracellular calcium levels. Hypoosmotic shock-induced cell swelling and concomitant calcium influx were reduced in the pkd2 mutants, suggesting that Pkd2 might act as a force-sensitive ion channel. In vitro reconstitution of the Pkd2 channel carried out with my collaborator Yen-Yu in the group of Allen Liu, discovered that Pkd2 is activated by membrane stretching and is permeable to calcium. In vivo, mutations of pkd2 also reduced the calcium spikes during cell separation, suggesting the role of this channel during cytokinesis. We concluded that Pkd2 allows calcium influx during cytokinesis when activated by membrane stretching force, likely representing a force-sensitive channel.

Autosomal Dominant Polycystic Kidney Disease a monogenic inherited human disease causing renal epithelial cells to proliferate forming fluid-filled cysts resulting in renal failure. Currently, no treatment exists to prevent or slower cyst formation, and most ADPKD patients require renal transplantation or life-long hemodialysis for survival. Mutations in either PKD1 or PKD2, are the underlying cause of ADPKD, with PKD1 mutations accounting for over 85% of the cases. Polycystin-1 (PC1) is an integral membrane glycoprotein containing 4302 amino acids with putative extracytoplasmic ligand-binding domains, but no physiological ligand has been identified to date. It has 11 transmembrane domains with an extracytoplasmic region of ~3000 amino acids and a cytoplasmic tail of ~200 amino acids. Many signaling pathways has been implicated to play a causative role for ADPKD but none show a clear mechanism. The study of STAT6, P100 and the PC1 tail using reporter assays, binding experiments, and immunohistochemistry suggested a model where the PC1 tail is cleaved due to loss of fluid flow, which triggers the nuclear translocation the constitutive trimeric complex comprising of the cleaved PC1 tail, STAT6 and P100 in renal epithelial cells. Introduction of C-terminus and the complete PC1 tail mRNA in zebra fish embryos resulted in cyst formation in their pronephric duct. Increased STAT6, P100 and PC1 tail staining in ADPKD kidney tissue sections also proved the importance of this pathway in stimulating cystogenesis. The membrane anchored PC1 tail was sufficient to activate constitutive pY-STAT3 and whereas the cleaved PC1 tail stimulated IFN-gamma stimulated reporter. This brings out a model where the PC1 can act as a molecular switch whereby it can regulate different STAT pathways depending on cellular conditions. Constitutive activation of STAT3 by normal PC1 may suggest the active STAT pathway under normal condition. Cleavage of the PC1 tail due to renal injury changes the function of the tail as a transcriptional co-activator which may now either activate STAT6 which would lead to proliferation or STAT1 or STAT3 leading to apoptosis. Inhibitors of the pathways that are activated in ADPKD can prove to be potential therapies for the disease.