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Desensitization of Inositol 1,4,5-Trisphosphate/Ca2+-induced Cl- Currents by Prolonged Activation of G Proteins in Xenopus Oocytes

Quick, Michael W. and Lester, Henry A. and Davidson, Norman and Simon, Melvin I. and Aragay, Anna M. (1996) Desensitization of Inositol 1,4,5-Trisphosphate/Ca2+-induced Cl- Currents by Prolonged Activation of G Proteins in Xenopus Oocytes. Journal of Biological Chemistry, 271 (50). pp. 32021-32027. ISSN 0021-9258.

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Expression of G protein alpha subunits of the Gq family with various G protein-coupled receptors induces activation of an inositol 1,4,5-trisphosphate (IP3)/Ca2+-mediated Cl- conductance in Xenopus oocytes. Our present data show that two members of this family, the human Galpha 16 subunit and the murine homologue Galpha 15, can induce both activation and inhibition of these agonist-induced currents. Although extremely low amounts (10-50 pg) of injected Galpha 16 subunit cRNA cause modest (~2-fold) enhancement of ligand-induced Cl- currents in oocytes co-injected with thyrotropin-releasing hormone (TRH) receptor cRNA 48 h postinjection, larger Galpha 16 and Galpha 15 cRNA injections cause >10-fold inhibition of TRH or 5HT2c receptor responses. The inhibition is analyzed in this study. The inhibited currents are recovered if various Gbeta gamma subunit combinations are also expressed with the Galpha subunits. The constitutively active mutant, Galpha 16Q212L, also causes a strong attenuation of the ligand-induced Cl- currents, but this inhibition is not recovered by co-expression of Gbeta gamma subunits. These results indicate that the free Galpha subunit is responsible for the inhibitory signal. Although expression of TRH receptor alone produces maximum responses approximately 48 h after injection, co-expression of TRH receptor with Galpha 16 results in enhanced responses 6-12 h postinjection, followed by complete attenuation at 36 h. Furthermore, injection of Galpha 16 cRNA alone at comparable levels gives rise to spontaneous Cl- currents within 6-12 h postinjection, suggesting that the early spontaneous activation underlies the later suppression. Expression of other G protein alpha subunits of the Gq family, at cRNA levels considerably higher than effective for Galpha 16, produces both analogous spontaneous Cl- currents and, later, inhibition of ligand-induced Cl- currents. Experiments with direct injection of IP3 and of Ca2+ suggest that this inhibition is consistent with the down-regulation of IP3 receptors. These data indicate that both enhancement and inhibition of signaling through G protein-coupled receptors can be mediated by the expression level and/or activity of an individual G protein.

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Lester, Henry A.0000-0002-5470-5255
Additional Information:© 1996 by The American Society for Biochemistry and Molecular Biology, Inc. (Received for publication, June 26, 1996, and in revised form, September 12, 1996) We thank Brad Henkle for oocyte preparation and Lorna Brundage for critical reading of the manuscript. We are also grateful to Dr. T. Wilkie for providing the Galpha 15Q121L cDNA. This work was supported in part by W. M. Keck Foundation Grant 931360 (to M. W. Q.) and National Institute of Health Grants CA13148 (to M. W. Q.) and MH-49176 (to M. I. S. and H. A. L.). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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W. M. Keck Foundation931360
Issue or Number:50
Record Number:CaltechAUTHORS:QUIjbc96
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ID Code:4668
Deposited By: Archive Administrator
Deposited On:02 Sep 2006
Last Modified:02 Oct 2019 23:15

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