RGS protein specificity towards Gq- and Gi/o-mediated ERK 1/2 and Akt activation, in vitro.
Extracellular Regulated Kinases (ERK) and Protein Kinase B (Akt) are intermediaries in relaying extracellular growth signals to intracellular targets. Each pathway can become activated upon stimulation of G protein-coupled receptors mediated by G(q) and G(i/o) proteins subjected to regulation by RGS proteins. The goal of the study was to delineate the specificity in which cardiac RGS proteins modulate G(q)and G(i/o)-induced ERK and Akt phosphorylation. To isolate G(q)- and G(i/o)-mediated effects, we exclusively expressed muscarinic M(2) or M(3) receptors in COS-7 cells. Western blot analyses demonstrated increase of phosphorylation of ERK 1.7-/3.3-fold and Akt 2.4-/6-fold in M(2)-/M(3)- expressing cells through carbachol stimulation. In co-expressions, M(3)/G(q)-induced activation of Akt was exclusively blunted through RGS3s/RGS3, whereas activation of ERK was inhibited additionally through RGS2/RGS5. M(2)/G(i/o) induced Akt activation was inhibited by all RGS proteins tested. RGS2 had no effect on M(2)/G(i/o)-induced ERK activation. The high degree of specificity in RGS proteins-depending modulation of G(q)- and G(i/o)-mediated ERK and Akt activation in the muscarinic network cannot merely be attributed exclusively to RGS protein selectivity towards G(q) or G(i/o) proteins. Counter-regulatory mechanisms and inter-signaling cross-talk may alter the sensitivity of GPCR-induced ERK and Akt activation to RGS protein regulation.
Department for Cardiology, Friedrich-Alexander University Erlangen, 91054 Erlangen, German. th_anger@arcor.de
Thomas
T
Anger
Nils
N
Klintworth
Christian
C
Stumpf
Werner G
WG
Daniel
Ulrike
U
Mende
Christoph D
CD
Garlichs
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D032383:Myocytes, Cardiac / Q000378:metabolism; D054799:Phospholipase C beta / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D020710:RGS Proteins / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D020239:Receptor Cross-Talk; D043585:Receptor, Muscarinic M2 / Q000235:genetics / Q000378:metabolism; D043587:Receptor, Muscarinic M3 / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection
D020710:RGS Proteins; D043585:Receptor, Muscarinic M2; D043587:Receptor, Muscarinic M3; D011994:Recombinant Proteins; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D054799:Phospholipase C beta; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D043802:GTP-Binding Protein alpha Subunits, Gq-G11