Regulatory Roles of Zinc Fluxes in Metalloprotein Occupancy and Cell Cycle Progression

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 DESCRIPTION (provided by applicant): Zinc fluxes have recently been discovered to be key regulatory events in a variety of human physiologies. For instance, significant zinc mobilization events play essential roles cortical neuron function, proper immune response, and insulin secretion. Zinc has traditionally been viewed as a static cofactor involved in protein structure and enzyme catalysis. More recently, however, a number of studies have provided support for the idea that zinc binding sites in regulatory proteins respond to transient fluctuations in zinc availability and are switched on and off in way that regulates key cellular events. Despite its rol in these profound physiologies the biochemistry of zinc action is not well understood. Little is known regarding the instructive mechanisms of zinc fluxes, zinc trafficking pathways and the role of specific metalloprotein receptors in signaling events. To interpret and eventually intervene in neurological disorders and metabolic diseases caused by disruption of such pathways, we plan to elucidate the basic scientific mechanisms of zinc-dependent switching events at the level of molecules and cells. We will test the hypothesis that regulatory zinc fluxes exert instructive control of the mammalian cell cycle through specific, receptor-mediated processes. This hypothesis is based on multiple lines of evidence, including: (1) data showing fluctuations zinc distribution at various points in the cell cycle for single cells; (2) live cell imaging demonstrating the movement of waves of zinc; and (3) physiochemical approaches showing colocalization of zinc with specific factors. We will use mammalian gametes, i.e. the sperm and egg, as a model cellular system to understand how an essential zinc signaling pathway works. One objective is to identify molecular mediators and targets of regulatory zinc fluxes by examining the quantitative changes in the localization of zinc in single cells at specifi points in the cell cycle. We will then correlate these spatio- temporal characteristics of metal availability with changes in protein chemistry of zinc receptors that are targets of the signaling events. Refinement and validation of the new chemical probes and physical methods developed in these studies will substantially enhance our knowledge of a fundamental pathway that regulates cellular decision making processes. At the project's conclusion, we will have accomplished three innovative goals of significance to the biomedical community. First, we will provide fundamental new insights into inorganic signaling events work at a molecular level. Second, we will develop methods to probe the metalloproteome and identify how key proteins involved in embryogenesis change zinc occupancy immediately following fertilization. Third, we will provide fundamental new insights into inorganic signaling events in sperm capacitation and acrosome reactions. Taken together these results will elucidate a robust role for proteins involved in zinc signaling fluxes and the mechanisms of cell cycle regulation.