Recruitment of RGS2 and RGS4 to the plasma membrane by G proteins and receptors reflects functional interactions.
N-terminally green fluorescent protein (GFP)-tagged regulator of G protein signaling (RGS) 2 and RGS4 fusion proteins expressed in human embryonic kidney 293 cells localized to the nucleus and cytosol, respectively. They were selectively recruited to the plasma membrane by G proteins and correspondingly by receptors that activate those G proteins: GFP-RGS2 when coexpressed with Galphas, beta2-adrenergic receptor, Galphaq, or AT1A angiotensin II receptor, and GFP-RGS4 when coexpressed with Galphai2 or M2 muscarinic receptor. G protein mutants with reduced RGS affinity did not produce this effect, implying that the recruitment involves direct binding to G proteins and is independent of downstream signaling events. Neither agonists nor inverse agonists altered receptor-promoted RGS association with the plasma membrane, and expressing either constitutively activated or poorly activated G protein mutants produced effects similar to those of their wild-type counterparts. Thus, intracellular interactions between these proteins seem to be relatively stable and insensitive to the activation state of the G protein, in contrast to the transient increases in RGS-G protein association known to be caused by G protein activation in solution-based assays. G protein effects on RGS localization were mirrored by RGS effects on G protein function. RGS4 was more potent than RGS2 in promoting steady-state Gi GTPase activity, whereas RGS2 inhibited Gs-dependent increases in intracellular cAMP, suggesting that G protein signaling in cells is regulated by the selective recruitment of RGS proteins to the plasma membrane.
Roy
Anju Anne
AA
Department of Physiology and Pharmacology, Medical Sciences Building, University of Western Ontario, London, Ontario N6A 5C1, Canada.
Lemberg
Kara E
KE
Chidiac
Peter
P
D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism / Q000502:physiology; D002522:Chlorocebus aethiops; D004305:Dose-Response Relationship, Drug; D019204:GTP-Binding Proteins / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D007424:Intracellular Fluid / Q000378:metabolism / Q000502:physiology; D021381:Protein Transport; D020710:RGS Proteins / Q000378:metabolism* / Q000502:physiology; D011956:Receptors, Cell Surface / Q000819:agonists / Q000378:metabolism* / Q000502:physiology
D020710:RGS Proteins; C087182:RGS2 protein, human; D011956:Receptors, Cell Surface; C098799:RGS4 protein; D019204:GTP-Binding Proteins