title: Spinophilin regulates Ca2+ signalling by binding the N-terminal domain of RGS2 and the third intracellular loop of G-protein-coupled receptors.
abstract: Signalling by G proteins is controlled by the regulator of G-protein signalling (RGS) proteins that accelerate the GTPase activity of Galpha subunits and act in a G-protein-coupled receptor (GPCR)-specific manner. The conserved RGS domain accelerates the G subunit GTPase activity, whereas the variable amino-terminal domain participates in GPCR recognition. How receptor recognition is achieved is not known. Here, we show that the scaffold protein spinophilin (SPL), which binds the third intracellular loop (3iL) of several GPCRs, binds the N-terminal domain of RGS2. SPL also binds RGS1, RGS4, RGS16 and GAIP. When expressed in Xenopus laevis oocytes, SPL markedly increased inhibition of alpha-adrenergic receptor (alphaAR) Ca2+ signalling by RGS2. Notably, the constitutively active mutant alphaAR(A293E) (the mutation being in the 3iL) did not bind SPL and was relatively resistant to inhibition by RGS2. Use of betaAR-alphaAR chimaeras identified the 288REKKAA293 sequence as essential for the binding of SPL and inhibition of Ca2+ signalling by RGS2. Furthermore, alphaAR-evoked Ca2+ signalling is less sensitive to inhibition by SPL in rgs2-/- cells and less sensitive to inhibition by RGS2 in spl-/- cells. These findings provide a general mechanism by which RGS proteins recognize GPCRs to confer signalling specificity.
authors.affiliation: Department of Physiology, University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390-9040, USA.
authors.forename: Xinhua
authors.initials: X
authors.lastname: Wang
authors.forename: Weizhong
authors.initials: W
authors.lastname: Zeng
authors.forename: Abigail A
authors.initials: AA
authors.lastname: Soyombo
authors.forename: Wei
authors.initials: W
authors.lastname: Tang
authors.forename: Elliott M
authors.initials: EM
authors.lastname: Ross
authors.forename: Anthony P
authors.initials: AP
authors.lastname: Barnes
authors.forename: Sharon L
authors.initials: SL
authors.lastname: Milgram
authors.forename: Josef M
authors.initials: JM
authors.lastname: Penninger
authors.forename: Patrick B
authors.initials: PB
authors.lastname: Allen
authors.forename: Paul
authors.initials: P
authors.lastname: Greengard
authors.forename: Shmuel
authors.initials: S
authors.lastname: Muallem
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D051379:Mice; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D009865:Oocytes / Q000737:chemistry; D011485:Protein Binding / Q000502:physiology; D020710:RGS Proteins / Q000378:metabolism / Q000502:physiology*; D043562:Receptors, G-Protein-Coupled / Q000187:drug effects / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D014982:Xenopus laevis; D000099273:Neurabins
chemical_list: D002118:Calcium; D008840:Microfilament Proteins; D009419:Nerve Tissue Proteins; D020710:RGS Proteins; D043562:Receptors, G-Protein-Coupled; D000099273:Neurabins