Differential effects of the Gβ5-RGS7 complex on muscarinic M3 receptor-induced Ca2+ influx and release

Darla Karpinsky-Semper, Claude Henry Volmar, Shaun P Brothers, Vladlen Z Slepak

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

The G protein β subunit Gβ5 uniquely forms heterodimers with R7 family regulators of G protein signaling (RGS) proteins (RGS6, RGS7, RGS9, and RGS11) instead of Gγ. Although the Gβ5-RGS7 complex attenuates Ca2+ signaling mediated by the muscarinic M3 receptor (M3R), the route of Ca2+entry (i.e., release from intracellular stores and/or influx across the plasma membrane) is unknown. Here, we show that, in addition to suppressing carbachol-stimulated Ca2+ release, Gβ5-RGS7 enhanced Ca2+ influx. This novel effect of Gβ5-RGS7 was blocked by nifedipine and 2-aminoethoxydiphenyl borate. Experiments with pertussis toxin, an RGS domain-deficient mutant of RGS7, and UBO-QIC {L-threonine, (3R)-N-acetyl-3-hydroxy-L-leucyl-(aR)-a-hydroxybenzenepropanoyl- 2,3-idehydro-N-methylalanyl-L-alanyl-N-methyl-L-alanyl- (3R)-3-[[(2S,3R)-3- hydroxy-4- methyl-1-oxo-2-[(1-oxopropyl)amino] pentyl]oxy]-L-leucyl-N,O- dimethyl-,(7→1)-lactone (9CI)}, a novel inhibitor of Gq, showed that Gβ5-RGS7 modulated a Gq-mediated pathway. These studies indicate that Gβ5-RGS7, independent of RGS7 GTPase-accelerating protein activity, couples M3R to a nifedipine-sensitive Ca2+ channel. We also compared the action of Gβ5-RGS7 on M3R-induced Ca2+ influx and release elicited by different muscarinic agonists. Responses to Oxo-M [oxotremorine methiodide N,N,N,-trimethyl-4-(2-oxo-1-pyrrolidinyl)-2-butyn-1- ammonium iodide] were insensitive to Gβ5-RGS7. Pilocarpine responses consisted of a large release and modest influx components, of which the former was strongly inhibited whereas the latter was insensitive to Gβ5-RGS7. McN-A-343 [(4-hydroxy-2- butynyl)-1-trimethylammonium-3-chlorocarbanilate chloride] was the only compound whose total Ca2+ response was enhanced by Gβ5- RGS7, attributed to, in part, by the relatively small Ca2+ release this partial agonist stimulated. Together, these results show that distinct agonists not only have differential M3R functional selectivity, but also confer specific sensitivity to the Gβ5-RGS7 complex.

Original languageEnglish
Pages (from-to)758-768
Number of pages11
JournalMolecular Pharmacology
Volume85
Issue number5
DOIs
StatePublished - Jan 1 2014

Fingerprint

Muscarinic M3 Receptors
Nifedipine
GTP-Binding Proteins
(4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethylammonium Chloride
RGS Proteins
Muscarinic Agonists
Pilocarpine
GTP Phosphohydrolases
Pertussis Toxin
Protein Subunits
Carbachol
Lactones
Threonine
Chlorides
Cell Membrane
Proteins

ASJC Scopus subject areas

  • Pharmacology
  • Molecular Medicine

Cite this

Differential effects of the Gβ5-RGS7 complex on muscarinic M3 receptor-induced Ca2+ influx and release. / Karpinsky-Semper, Darla; Volmar, Claude Henry; Brothers, Shaun P; Slepak, Vladlen Z.

In: Molecular Pharmacology, Vol. 85, No. 5, 01.01.2014, p. 758-768.

Research output: Contribution to journalArticle

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AB - The G protein β subunit Gβ5 uniquely forms heterodimers with R7 family regulators of G protein signaling (RGS) proteins (RGS6, RGS7, RGS9, and RGS11) instead of Gγ. Although the Gβ5-RGS7 complex attenuates Ca2+ signaling mediated by the muscarinic M3 receptor (M3R), the route of Ca2+entry (i.e., release from intracellular stores and/or influx across the plasma membrane) is unknown. Here, we show that, in addition to suppressing carbachol-stimulated Ca2+ release, Gβ5-RGS7 enhanced Ca2+ influx. This novel effect of Gβ5-RGS7 was blocked by nifedipine and 2-aminoethoxydiphenyl borate. Experiments with pertussis toxin, an RGS domain-deficient mutant of RGS7, and UBO-QIC {L-threonine, (3R)-N-acetyl-3-hydroxy-L-leucyl-(aR)-a-hydroxybenzenepropanoyl- 2,3-idehydro-N-methylalanyl-L-alanyl-N-methyl-L-alanyl- (3R)-3-[[(2S,3R)-3- hydroxy-4- methyl-1-oxo-2-[(1-oxopropyl)amino] pentyl]oxy]-L-leucyl-N,O- dimethyl-,(7→1)-lactone (9CI)}, a novel inhibitor of Gq, showed that Gβ5-RGS7 modulated a Gq-mediated pathway. These studies indicate that Gβ5-RGS7, independent of RGS7 GTPase-accelerating protein activity, couples M3R to a nifedipine-sensitive Ca2+ channel. We also compared the action of Gβ5-RGS7 on M3R-induced Ca2+ influx and release elicited by different muscarinic agonists. Responses to Oxo-M [oxotremorine methiodide N,N,N,-trimethyl-4-(2-oxo-1-pyrrolidinyl)-2-butyn-1- ammonium iodide] were insensitive to Gβ5-RGS7. Pilocarpine responses consisted of a large release and modest influx components, of which the former was strongly inhibited whereas the latter was insensitive to Gβ5-RGS7. McN-A-343 [(4-hydroxy-2- butynyl)-1-trimethylammonium-3-chlorocarbanilate chloride] was the only compound whose total Ca2+ response was enhanced by Gβ5- RGS7, attributed to, in part, by the relatively small Ca2+ release this partial agonist stimulated. Together, these results show that distinct agonists not only have differential M3R functional selectivity, but also confer specific sensitivity to the Gβ5-RGS7 complex.

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