EP4Prostanoid Receptor Coupling to a Pertussis Toxin-Sensitive Inhibitory G Protein

We investigated the mechanisms of receptor-mediated stimulation of high-affinity GTPase activity in response to opioid peptides and to foetal-calf serum in membranes of the neuroblastoma X glioma hybrid cell line NG108-15. Increases in GTPase activity in response to both of these ligands was abolished by prior exposure of the cells to pertussis toxin. Pertussis toxin in the presence of [32P]NAD+ catalysed incorporation of radioactivity into a broad band of approx. 40 kDa in membranes prepared from untreated, but not from pertussis-toxin-pretreated, cells. Additivity studies indicated that the responses to opioid peptides and to foetal-calf serum were mediated by separate guanine-nucleotide-binding proteins (G-proteins). Whereas opioid peptides produced an inhibition of adenylate cyclase in membranes of untreated cells, foetal-calf serum did not. Affinity-purified antibodies which recognize the C-terminus of the inhibitory G-protein identified a 40 kDa polypeptide in membranes of NG108-15 cells. These antibodies attenuated opioid-stimulated high-affinity GTPase activity, but did not markedly affect the response to foetal-calf serum. We conclude that receptors for the opioid peptides function via the inhibitory G-protein (Gi), whereas foetal-calf serum activates a second pertussis-toxin-sensitive G-protein, which has a C-terminal sequence significantly different from that of Gi.

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Mechanisms of action of angiotensin II on mammalian sperm function

Reproduction ◽

10.1530/rep.1.00457 ◽

2005 ◽

Vol 129 (2) ◽

pp. 211-218 ◽

Cited By ~ 9

Author(s):

Samra Mededovic ◽

Lynn R Fraser

Keyword(s):

Angiotensin Ii ◽

Pertussis Toxin ◽

Fluorescence Analysis ◽

Camp Production ◽

Receptor Coupling ◽

Mammalian Sperm ◽

Significant Acceleration ◽

Intracellular Ca ◽

Inhibitory G Protein ◽

Stimulation Of

Angiotensin II (AII) stimulates capacitation and fertilizing ability in mammalian spermatozoa, with the binding of AII to its receptors resulting in stimulation of cAMP production in both uncapacitated and capacitated cells. This study investigated possible mechanisms whereby AII affects cAMP availability. The first question was whether extracellular Ca2+is required for responses in mouse spermatozoa and, using chlortetracycline fluorescence analysis, it was clear that cells responded to AII only when the medium contained CaCl2, with both 90 μM and 1.80 mM supporting a significant acceleration of capacitation. Consistent with those results, AII significantly stimulated cAMP production in both CaCl2-containing media tested, the response being greater in that containing 1.80 mM. Several different agents that might affect the signalling pathway stimulated by AII were then evaluated in uncapacitated suspensions. Chlortetracycline analysis revealed that pertussis toxin abolished responses to AII, suggesting the involvement of an inhibitory Gα subunit; dideoxyadenosine, a specific membrane-associated adenylyl cyclase (mAC) P-site inhibitor, also blocked responses, suggesting involvement of an mAC. cAMP determinations con-firmed that both reagents also abolished AII’s stimulation of cAMP. In contrast, nifedipine, a Ca2+channel blocker, did not inhibit AII’s effects on spermatozoa. Finally, in capacitated suspensions, both pertussis toxin and dideoxyadenosine were again shown to block AII’s stimulation of cAMP. These results suggest that responses to AII involve an inhibitory G protein and an mAC, but it is likely that AII–receptor coupling does not stimulate directly mAC but rather does so in an indirect manner, perhaps by altering the intracellular Ca2+concentration.

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THE EFFECTS OF PERTUSSIS TOXIN ON VASOCONSTRICTOR AND VASODILATOR AGENTS IN THE PITHED RAT MAY NOT BE AN INDICATOR OF G-PROTEIN RECEPTOR COUPLING

Clinical and Experimental Pharmacology and Physiology ◽

10.1111/j.1440-1681.1993.tb01665.x ◽

1993 ◽

Vol 20 (3) ◽

pp. 169-176

Author(s):

Mary Costa ◽

Henryk Majewski

Keyword(s):

G Protein ◽

Pertussis Toxin ◽

Pithed Rat ◽

Vasodilator Agents ◽

Receptor Coupling ◽

Protein Receptor

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Cardioprotective Trafficking of Caveolin to Mitochondria Is Gi-protein Dependent

Anesthesiology ◽

10.1097/aln.0000000000000295 ◽

2014 ◽

Vol 121 (3) ◽

pp. 538-548 ◽

Cited By ~ 21

Author(s):

Jiawan Wang ◽

Jan M. Schilling ◽

Ingrid R. Niesman ◽

John P. Headrick ◽

J. Cameron Finley ◽

...

Keyword(s):

G Protein ◽

Pertussis Toxin ◽

Glycogen Synthase ◽

Caveolin 1 ◽

Beneficial Effects ◽

Group Data ◽

Gi Protein ◽

Inhibitory G Protein ◽

Group Protein ◽

Protective Signaling

Abstract Background: Caveolae are a nexus for protective signaling. Trafficking of caveolin to mitochondria is essential for adaptation to cellular stress though the trafficking mechanisms remain unknown. The authors hypothesized that G protein–coupled receptor/inhibitory G protein (Gi) activation leads to caveolin trafficking to mitochondria. Methods: Mice were exposed to isoflurane or oxygen vehicle (30 min, ±36 h pertussis toxin pretreatment, an irreversible Gi inhibitor). Caveolin trafficking, cardioprotective “survival kinase” signaling, mitochondrial function, and ultrastructure were assessed. Results: Isoflurane increased cardiac caveolae (n = 8 per group; data presented as mean ± SD for Ctrl versus isoflurane; [caveolin-1: 1.78 ± 0.12 vs. 3.53 ± 0.77; P < 0.05]; [caveolin-3: 1.68 ± 0.29 vs. 2.67 ± 0.46; P < 0.05]) and mitochondrial caveolin levels (n = 16 per group; [caveolin-1: 0.87 ± 0.18 vs. 1.89 ± .19; P < 0.05]; [caveolin-3: 1.10 ± 0.29 vs. 2.26 ± 0.28; P < 0.05]), and caveolin-enriched mitochondria exhibited improved respiratory function (n = 4 per group; [state 3/complex I: 10.67 ± 1.54 vs. 37.6 ± 7.34; P < 0.05]; [state 3/complex II: 37.19 ± 4.61 vs. 71.48 ± 15.28; P < 0.05]). Isoflurane increased phosphorylation of survival kinases (n = 8 per group; [protein kinase B: 0.63 ± 0.20 vs. 1.47 ± 0.18; P < 0.05]; [glycogen synthase kinase 3β: 1.23 ± 0.20 vs. 2.35 ± 0.20; P < 0.05]). The beneficial effects were blocked by pertussis toxin. Conclusions: Gi proteins are involved in trafficking caveolin to mitochondria to enhance stress resistance. Agents that target Gi activation and caveolin trafficking may be viable cardioprotective agents.

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