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Journal of Pharmacology and Experimental Therapeutics

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Research ArticleCELLULAR AND MOLECULAR

Direct Interactions between the Heterotrimeric G Protein Subunit Gβ5 and the G Protein γ Subunit-Like Domain-Containing Regulator of G Protein Signaling 11: Gain of Function of Cyan Fluorescent Protein-Tagged Gγ3

Janice Y. Zhou, Peter T. Toth and Richard J. Miller
Journal of Pharmacology and Experimental Therapeutics May 2003, 305 (2) 460-466; DOI: https://doi.org/10.1124/jpet.102.048637
Janice Y. Zhou
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Peter T. Toth
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Richard J. Miller
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Abstract

We used fluorescence resonance energy transfer imaging of enhanced cyan fluorescent protein (CFP)-tagged and enhanced yellow fluorescent protein (YFP)-tagged protein pairs to examine the hypothesis that G protein γ subunit-like (GGL) domain-containing regulators of G protein signaling (RGS) can directly bind to the Gβ5 subunit of heterotrimeric G proteins in vivo. We observed that Gβ5 could interact with Gγ2 and Gγ13, after their expression in human embryonic kidney 293 cells. Interestingly, although untagged Gγ3 did not interact with Gβ5, CFP-tagged Gγ3 strongly interacted with YFP-tagged Gβ5 in FRET studies. Moreover, CFP-Gγ3 supported Ca2+ channel inhibition when paired with Gβ5 or YFP-Gβ5, indicating a “gain of function” for CFP-Gγ3. Gβ5 could also interact with RGS11 and its N-terminal, but not its C-terminal domain. On the other hand, RGS11 did not interact with Gβ1. These studies demonstrate that the GGL domain-containing N terminus of RGS 11 can directly interact with Gβ5 in vivo and supports the hypothesis that this interaction may contribute to the specificity of Gβ5 interactions with cellular effector molecules.

Footnotes

  • This study was supported by U.S. Public Health Service Grant MH-40165. Preliminary results were presented in abstract format at the 2001 Neuroscience Meeting (Abstract 271.7).

  • DOI: 10.1124/jpet.102.048637

  • Abbreviations:
    GPCR
    G protein-coupled receptor
    VDCC
    voltage-dependent Ca2+ channel
    RGS
    regulator of G protein signaling
    GGL
    G protein γ subunit-like
    GTPase-activating protein
    FRET, fluorescence resonance energy transfer
    YFP
    yellow fluorescent protein
    CFP
    cyan fluorescent protein
    ANOVA
    analysis of variance
    FRETN
    normalized fluorescence resonance energy transfer
    ROI
    region of interest
    aa
    amino acid
    • Received January 1, 2003.
    • Accepted February 6, 2003.
  • The American Society for Pharmacology and Experimental Therapeutics
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Journal of Pharmacology and Experimental Therapeutics: 305 (2)
Journal of Pharmacology and Experimental Therapeutics
Vol. 305, Issue 2
1 May 2003
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Research ArticleCELLULAR AND MOLECULAR

Direct Interactions between the Heterotrimeric G Protein Subunit Gβ5 and the G Protein γ Subunit-Like Domain-Containing Regulator of G Protein Signaling 11: Gain of Function of Cyan Fluorescent Protein-Tagged Gγ3

Janice Y. Zhou, Peter T. Toth and Richard J. Miller
Journal of Pharmacology and Experimental Therapeutics May 1, 2003, 305 (2) 460-466; DOI: https://doi.org/10.1124/jpet.102.048637

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Research ArticleCELLULAR AND MOLECULAR

Direct Interactions between the Heterotrimeric G Protein Subunit Gβ5 and the G Protein γ Subunit-Like Domain-Containing Regulator of G Protein Signaling 11: Gain of Function of Cyan Fluorescent Protein-Tagged Gγ3

Janice Y. Zhou, Peter T. Toth and Richard J. Miller
Journal of Pharmacology and Experimental Therapeutics May 1, 2003, 305 (2) 460-466; DOI: https://doi.org/10.1124/jpet.102.048637
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