Synergistic Ca2+ Responses by Gαi- and Gαq-coupled G-protein-coupled Receptors Require a Single PLCβ Isoform That Is Sensitive to Both Gβγ and Gαq

被引:48
作者
Rebres, Robert A. [1 ]
Roach, Tamara I. A. [1 ]
Fraser, Iain D. C. [3 ]
Philip, Finly [4 ]
Moon, Christina [1 ]
Lin, Keng-Mean [4 ]
Liu, Jamie [3 ]
Santat, Leah [3 ]
Cheadle, Lucas [3 ]
Ross, Elliott M. [4 ]
Simon, Melvin I. [1 ]
Seaman, William E. [2 ]
机构
[1] Univ Calif San Francisco, Vet Affairs Med Ctr, No Calif Inst Res & Educ, San Francisco, CA 94121 USA
[2] Univ Calif San Francisco, Vet Affairs Med Ctr, Dept Med, San Francisco, CA 94121 USA
[3] CALTECH, Div Biol, Pasadena, CA 91125 USA
[4] Univ Texas SW Med Ctr Dallas, Dept Pharmacol, Dallas, TX 75390 USA
基金
美国国家卫生研究院;
关键词
PHOSPHOLIPASE-C-BETA; ADENOSINE A(1) RECEPTORS; ALPHA-SUBUNIT; CROSS-TALK; MEDIATED ACTIVATION; CELLS; CALCIUM; DOMAIN; STIMULATION; IDENTIFICATION;
D O I
10.1074/jbc.M110.198200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Cross-talk between G alpha(i)- and G alpha(q)-linked G-protein-coupled receptors yields synergistic Ca2+ responses in a variety of cell types. Prior studies have shown that synergistic Ca2+ responses from macrophage G-protein-coupled receptors are primarily dependent on phospholipase C beta 3 (PLC beta 3), with a possible contribution of PLC beta 2, whereas signaling through PLC beta 4 interferes with synergy. We here show that synergy can be induced by the combination of G beta gamma and G alpha(q) activation of a single PLC beta isoform. Synergy was absent in macrophages lacking both PLC beta 2 and PLC beta 3, but it was fully reconstituted following transduction with PLC beta 3 alone. Mechanisms of PLC beta mediated synergy were further explored in NIH-3T3 cells, which express little if any PLC beta 2. RNAi-mediated knockdown of endogenous PLC beta s demonstrated that synergy in these cells was dependent on PLC beta 3, but PLC beta 1 and PLC beta 4 did not contribute, and overexpression of either isoform inhibited Ca2+ synergy. When synergy was blocked by RNAi of endogenous PLC beta 3, it could be reconstituted by expression of either human PLC beta 3 or mouse PLC beta 2. In contrast, it could not be reconstituted by human PLC beta 3 with a mutation of the Y box, which disrupted activation by G beta gamma, and it was only partially restored by human PLC beta 3 with a mutation of the C terminus, which partly disrupted activation by G alpha(q). Thus, both G beta gamma and G alpha(q) contribute to activation of PLC beta 3 in cells for Ca2+ synergy. We conclude that Ca2+ synergy between G alpha(i)-coupled and G alpha(q)-coupled receptors requires the direct action of both G beta gamma and G alpha(q) on PLC beta and is mediated primarily by PLC beta 3, although PLC beta 2 is also competent.
引用
收藏
页码:942 / 951
页数:10
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