Analysis of differential modulatory activities of GRK2 and GRK4 on Galphaq-coupled receptor signaling.
G-protein-coupled receptor kinases (GRK) contain a regulator of G-protein signaling (RGS)-like domain located at the N terminus (GRK-Nter) of their sequence. This domain is present in all the GRK subtypes, but the RGS-like domain of GRK2 was documented to be functionally active, as it is able to interact selectively with Galphaq (both in vitro and in cells) and to inhibit Galphaq-dependent signaling. In contrast GRK4, GRK5, and GRK6 are unable to interact with Galphaq. This article describes the methodology to investigate the modulatory activity of GRK2 and GRK4 on GPCR-stimulated Galphaq signaling. This analysis is essentially based on three types of experiments: (a) study of the effect of the GRK-Nter on GPCR-dependent signaling; (b) analysis of the binding of GRK-Nter to Galphaq in vitro; and (c) analysis of the interaction of GRK with Galphaq in cells.
INM Neuromed, IRCCS, Pozzilli and Dpt of Human Physiology, University of Rome La Sapienza.
Antonietta
A
Picascia
Loredana
L
Capobianco
Luisa
L
Iacovelli
Antonio
A
De Blasi
D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D054771:G-Protein-Coupled Receptor Kinase 4; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D005810:Multigene Family; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D044405:Receptor, Serotonin, 5-HT2C / Q000235:genetics / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D018094:Receptors, Metabotropic Glutamate / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015290:Second Messenger Systems / Q000502:physiology*; D051552:beta-Adrenergic Receptor Kinases
D007527:Isoenzymes; D044405:Receptor, Serotonin, 5-HT2C; D043562:Receptors, G-Protein-Coupled; D018094:Receptors, Metabotropic Glutamate; D011993:Recombinant Fusion Proteins; D017346:Protein Serine-Threonine Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D051552:beta-Adrenergic Receptor Kinases; D054771:G-Protein-Coupled Receptor Kinase 4; C517855:GRK4 protein, human; D043802:GTP-Binding Protein alpha Subunits, Gq-G11