A quantitative characterization of the yeast heterotrimeric G protein cycle.
The yeast mating response is one of the best understood heterotrimeric G protein signaling pathways. Yet, most descriptions of this system have been qualitative. We have quantitatively characterized the heterotrimeric G protein cycle in yeast based on direct in vivo measurements. We used fluorescence resonance energy transfer to monitor the association state of cyan fluorescent protein (CFP)-Galpha and Gbetagamma-yellow fluorescent protein (YFP), and we found that receptor-mediated G protein activation produced a loss of fluorescence resonance energy transfer. Quantitative time course and dose-response data were obtained for both wild-type and mutant cells possessing an altered pheromone response. These results paint a quantitative portrait of how regulators such as Sst2p and the C-terminal tail of alpha-factor receptor modulate the kinetics and sensitivity of G protein signaling. We have explored critical features of the dynamics including the rapid rise and subsequent decline of active G proteins during the early response, and the relationship between the G protein activation dose-response curve and the downstream dose-response curves for cell-cycle arrest and transcriptional induction. Fitting the data to a mathematical model produced estimates of the in vivo rates of heterotrimeric G protein activation and deactivation in yeast.
Yi
Tau-Mu
TM
Systems Biology Group, Exploratory Research for Advanced Technology Kitano Symbiotic Systems Project, Japan Science and Technology Corporation, Shibuya, Tokyo, Japan.
Kitano
Hiroaki
H
Simon
Melvin I
MI
D004735:Energy Transfer; D005453:Fluorescence; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism*; D007700:Kinetics; D000072235:Mating Factor; D010455:Peptides / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D015398:Signal Transduction
D010455:Peptides; D000072235:Mating Factor; D020962:Heterotrimeric GTP-Binding Proteins