S-Nitrosothiols modulate G protein-coupled receptor signaling in a reversible and highly receptor-specific mannerReport as inadecuate




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BMC Cell Biology

, 6:21

First Online: 25 April 2005Received: 05 November 2004Accepted: 25 April 2005

Abstract

BackgroundRecent studies indicate that the G protein-coupled receptor GPCR signaling machinery can serve as a direct target of reactive oxygen species, including nitric oxide NO and S-nitrosothiols RSNOs. To gain a broader view into the way that receptor-dependent G protein activation – an early step in signal transduction – might be affected by RSNOs, we have studied several receptors coupling to the Gi family of G proteins in their native cellular environment using the powerful functional approach of SGTPγS autoradiography with brain cryostat sections in combination with classical G protein activation assays.

ResultsWe demonstrate that RSNOs, like S-nitrosoglutathione GSNO and S-nitrosocysteine CysNO, can modulate GPCR signaling via reversible, thiol-sensitive mechanisms probably involving S-nitrosylation. RSNOs are capable of very targeted regulation, as they potentiate the signaling of some receptors exemplified by the M2-M4 muscarinic cholinergic receptors, inhibit others P2Y12 purinergic, LPA1lysophosphatidic acid, and cannabinoid CB1 receptors, but may only marginally affect signaling of others, such as adenosine A1, μ-opioid, and opiate related receptors. Amplification of M2-M4 muscarinic responses is explained by an accelerated rate of guanine nucleotide exchange, as well as an increased number of high-affinity SGTPγS binding sites available for the agonist-activated receptor. GSNO amplified human M4 receptor signaling also under heterologous expression in CHO cells, but the effect diminished with increasing constitutive receptor activity. RSNOs markedly inhibited P2Y12 receptor signaling in native tissues rat brain and human platelets, but failed to affect human P2Y12 receptor signaling under heterologous expression in CHO cells, indicating that the native cellular signaling partners, rather than the P2Y12 receptor protein, act as a molecular target for this action.

ConclusionThese in vitro studies show for the first time in a broader general context that RSNOs are capable of modulating GPCR signaling in a reversible and highly receptor-specific manner. Given that the enzymatic machinery responsible for endogenous NO production is located in close proximity with the GPCR signaling complex, especially with that for several receptors whose signaling is shown here to be modulated by exogenous RSNOs, our data suggest that GPCR signaling in vivo is likely to be subject to substantial, and highly receptor-specific modulation by NO-derived RSNOs.

List of abbreviations2MeSADP2-methylthio-ADP

SGTPγSguanosine-5-O-3-S-thio-triphosphate

5-HT5-hydroxytryptamine

ADAadenosine deaminase

AGSactivator of G protein signaling

CChcarbacholine

CHOChinese hamster ovary

CP-55940-3-2-hydroxy-4-1,1-dimethylheptyl-phenyl-4-3-hydroxypropylcyclohexan-1-ol

CysNOS-nitrosocysteine

CysNO-GlyS-nitroso-cysteinyl-glycine

DAMGOD-Ala, N-Me-Phe, Gly-ol-enkephalin

DPCPX8-cyclopentyl-1,3-dipropylxanthine

DTTdithiotreitol

Glu-CysNOL-γ-glutamyl-S-nitrosocysteine

GSHglutathione

GPCRsG protein-coupled receptors

GSNOS-nitrosoglutathione

HAhemagglutinin

hM4human muscarinic receptor subtype 4

hP2Y12human P2Y12 purinergic receptor

LPAlysophosphatidic acid

NAnoradrenaline

NOnitric oxide

NOBF4nitrosodium tetrafluoroborate

NOSNO synthase

RGSregulator of G protein signaling

RTreverse transcriptase

SNAPS-nitroso-N-acetyl-D,L-penicillamine

RSNOS-nitrosothiol

SNPsodium nitroprusside

Electronic supplementary materialThe online version of this article doi:10.1186-1471-2121-6-21 contains supplementary material, which is available to authorized users.

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Author: Tarja Kokkola - Juha R Savinainen - Kati S Mönkkönen - Montse Durán Retamal - Jarmo T Laitinen

Source: https://link.springer.com/article/10.1186/1471-2121-6-21



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