REGULATING GLI FUNCTION IN HAIR FOLLICLE PROGENITORS

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DESCRIPTION (provided by applicant): Adult stem cells are multipotent cells that possess the capacity for programmed organ replacement and carry the promise of induced organ repair in response to injury or damage. Murine hair follicles provide an invaluable approach to modeling fundamental processes of regeneration because they undergo repeated, genetically controlled cycles of proliferation (anagen), regression (catagen) and quiescence (telogen). Intense recent focus has come to elucidate the controls of stem cell quiescence and the timing of re-entering the proliferative anagen stage. Previous studies have shown several key signaling pathways such as the Wnt and Sonic hedgehog (Shh) stimulate anagen, and studies funded through the parent grant 5ARO54780 have focused on the epithelial signaling inputs that control anagen via Shh signaling. By contrast, while recent data supports the calcineurin/NFAT pathways in maintaining quiescence, the higher order mechanisms of cycle timing remain poorly understood. The involvement of calcineurin/NFAT led us to investigate the role of calcium regulatory proteins in hair cycling. A screen for small molecules that topically regulate hair cycling revealed that nifedipine (DHP), an L-type voltage gated calcium channel antagonist, causes precocious entry to anagen in mice. Consistent with this finding, conditional ablation of CACNA1C (CaV1.2), the dominant cutaneous channel, causes similar hair cycling defects and precocious entry to anagen in mice. Further, mice containing a CaV1.2 gain-of-function mutation that mimics the Timothy syndrome (TS), a human multi-organ system disorder, delays entry into anagen. These data support the hypothesis that calcium transients via the voltage gated calcium channels regulate hair stem cell quiescence through Nfatc1 regulation. In order to further dissect the mechanism of calcium signaling and stem cell quiescence, we propose to: Dissect the molecular defects in of CaV1.2 mutant mice, examining the cell lineage requiring CaV1.2 function and the alterations in known signaling pathways in mutant mice; Measure the differences in amount of calcium in stem cells from wild type and Cav1.2 mutant mice in vitro and in vivo by using ratiometric calcium dyes and genetically-encoded calcium sensors. These studies will provide new insights into the timing mechanisms used during organogenesis. PUBLIC HEALTH RELEVENCE: Adult stem cells are multipotent cells that possess the capacity for programmed organ replacement and carry the promise of induced organ repair in response to injury or damage. We have found using small molecule antagonists and mouse mutants that L-type voltage gated calcium channels regulate the timing of hair regeneration. We believe that calcium transients via the voltage gated calcium channels regulate hair stem cell quiescence through Nfatc1 regulation. We propose to dissect the molecular defects in of CaV1.2 mutant mice, and measure the differences in amount of calcium in stem cells from wild type and Cav1.2 mutants. These studies will provide new insights into the timing mechanisms used during organogenesis.