期刊
SCIENCE SIGNALING
卷 3, 期 141, 页码 -出版社
AMER ASSOC ADVANCEMENT SCIENCE
DOI: 10.1126/scisignal.2000980
关键词
-
资金
- National Institute of Allergy and Infectious Diseases
- NIH
- American Heart Association [AHA0930127N]
The coupling of heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptors (GPCRs) with G proteins is fundamental for GPCR signaling; however, the mechanism of coupling is still debated. Moreover, how the proposed mechanisms affect the dynamics of downstream signaling remains unclear. Here, through experiments involving fluorescence recovery after photobleaching and single-molecule imaging, we directly measured the mobilities of cyclic adenosine monophosphate (cAMP) receptor 1 (cAR1), a chemoattractant receptor, and a G protein beta gamma subunit in live cells. We found that cAR1 diffused more slowly in the plasma membrane than did G beta gamma. Upon binding of ligand to the receptor, the mobility of cAR1 was unchanged, whereas the speed of a fraction of the faster-moving G beta gamma subunits decreased. Our measurements showed that cAR1 was relatively immobile and G beta gamma diffused freely, suggesting that chemoattractant-bound cAR1 transiently interacted with G proteins. Using models of possible coupling mechanisms, we computed the temporal kinetics of G protein activation. Our fluorescence resonance energy transfer imaging data showed that fully activated cAR1 induced the sustained dissociation of G protein alpha and beta gamma subunits, which indicated that ligand-bound cAR1 activated G proteins continuously. Finally, simulations indicated that a high-affinity coupling of ligand-bound receptors and G proteins was essential for cAR1 to translate extracellular gradient signals into directional cellular responses. We suggest that chemoattractant receptors use a ligand-induced coupling rather than a precoupled mechanism to control the activation of G proteins during chemotaxis.
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