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Science 291 (5512): 2408-2411

Copyright © 2001 by the American Association for the Advancement of Science

Receptor-Mediated Activation of Heterotrimeric G-Proteins in Living Cells

Chris Janetopoulos,* Tian Jin,* Peter Devreotes*dagger

Receptor-mediated activation of heterotrimeric GTP-binding proteins (G-proteins) was visualized in living Dictyostelium discoideum cells by monitoring fluorescence resonance energy transfer (FRET) between alpha - and beta - subunits fused to cyan and yellow fluorescent proteins. The G-protein heterotrimer rapidly dissociated and reassociated upon addition and removal of chemoattractant. During continuous stimulation, G-protein activation reached a dose-dependent steady-state level. Even though physiological responses subsided, the activation did not decline. Thus, adaptation occurs at another point in the signaling pathway, and occupied receptors, whether or not they are phosphorylated, catalyze the G-protein cycle. Construction of similar energy-transfer pairs of mammalian G-proteins should enable direct in situ mechanistic studies and applications such as drug screening and identifying ligands of newly found G-protein-coupled receptors.

Department of Biological Chemistry, Johns Hopkins Medical Institutions, Baltimore, MD 21205, USA.
*   Present address: Department of Cell Biology and Anatomy, Johns Hopkins Medical Institutions, Baltimore, MD 21205, USA.

dagger    To whom correspondence should be addressed.

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   Abstract »    Full Text »    PDF »
The CC Chemokine Eotaxin (CCL11) Is a Partial Agonist of CC Chemokine Receptor 2b.
R. Martinelli, I. Sabroe, G. LaRosa, T. J. Williams, and J. E. Pease (2001)
J. Biol. Chem. 276, 42957-42964
   Abstract »    Full Text »    PDF »
Allurin, a 21-kDa sperm chemoattractant from Xenopus egg jelly, is related to mammalian sperm-binding proteins.
J. H. Olson, X. Xiang, T. Ziegert, A. Kittelson, A. Rawls, A. L. Bieber, and D. E. Chandler (2001)
PNAS 98, 11205-11210
   Abstract »    Full Text »    PDF »
Reducing the Environmental Sensitivity of Yellow Fluorescent Protein: MECHANISM AND APPLICATIONS.
O. Griesbeck, G. S. Baird, R. E. Campbell, D. A. Zacharias, and R. Y. Tsien (2001)
J. Biol. Chem. 276, 29188-29194
   Abstract »    Full Text »    PDF »
Reducing the Environmental Sensitivity of Yellow Fluorescent Protein: MECHANISM AND APPLICATIONS.
O. Griesbeck, G. S. Baird, R. E. Campbell, D. A. Zacharias, and R. Y. Tsien (2001)
J. Biol. Chem. 276, 29188-29194
   Abstract »    Full Text »    PDF »
Kinetic Analysis of M2 Muscarinic Receptor Activation of Gi in Sf9 Insect Cell Membranes.
V. A. Mosser, I. J. Amana, and M. I. Schimerlik (2002)
J. Biol. Chem. 277, 922-931
   Abstract »    Full Text »    PDF »
Allurin, a 21-kDa sperm chemoattractant from Xenopus egg jelly, is related to mammalian sperm-binding proteins.
J. H. Olson, X. Xiang, T. Ziegert, A. Kittelson, A. Rawls, A. L. Bieber, and D. E. Chandler (2001)
PNAS 98, 11205-11210
   Abstract »    Full Text »    PDF »

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