QUANTITATIVE ANALYSIS OF G-PROTEINS & COUPLED RECEPTORS IN YEAST MATING

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. G-proteins, their coupled receptors, and underlying pathways are of great interest as targets for pharmaceutical development, with current levels of all approved therapeutic agents affecting these systems at greater than 50%. Using florescence techniques a greater understanding how G-proteins respond to extra cellular signals, mount a response, and the underlying regulation of such systems, which are highly conserved, contributes to the robustness and dynamic range of the sensors.Two-photon microscopy will be utilized to quantitate the location and dynamics of fluorescently labeled proteins, such as the receptor, g-proteins and downstream effectors. FRET, and FCS/ICS will be used to understand the molecular dynamics of the system, protein number, diffusion rate, and diffusion vector, and build a body of data that can then be used to make computational models. Other aspects of the mating process will also be observed, such as membrane fluidity and composition and it changes over time, localization of protein recruitment and internalization, and co-localization of proteins with these phenomena.