RGSZ1, a Gz-selective RGS protein in brain. Structure, membrane association, regulation by Galphaz phosphorylation, and relationship to a Gz gtpase-activating protein subfamily.
We cloned the cDNA for human RGSZ1, the major Gz-selective GTPase-activating protein (GAP) in brain (Wang, J., Tu, Y., Woodson, J., Song, X., and Ross, E. M. (1997) J. Biol. Chem. 272, 5732-5740) and a member of the RGS family of G protein GAPs. Its sequence is 83% identical to RET-RGS1 (except its N-terminal extension) and 56% identical to GAIP. Purified, recombinant RGSZ1, RET-RGS1, and GAIP each accelerated the hydrolysis of Galphaz-GTP over 400-fold with Km values of approximately 2 nM. RGSZ1 was 100-fold selective for Galphaz over Galphai, unusually specific among RGS proteins. Other enzymological properties of RGSZ1, brain Gz GAP, and RET-RGS1 were identical; GAIP differed only in Mg2+ dependence and in its slightly lower selectivity for Galphaz. RGSZ1, RET-RGS1, and GAIP thus define a subfamily of Gz GAPs within the RGS proteins. RGSZ1 has no obvious membrane-spanning region but is tightly membrane-bound in brain. Its regulatory activity in membranes depends on stable bilayer association. When co-reconstituted into phospholipid vesicles with Gz and m2 muscarinic receptors, RGSZ1 increased agonist-stimulated GTPase >15-fold with EC50 <12 nM, but RGSZ1 added to the vesicle suspension was <0.1% as active. RGSZ1, RET-RGS1, and GAIP share a cysteine string sequence, perhaps targeting them to secretory vesicles and allowing them to participate in the proposed control of secretion by Gz. Phosphorylation of Galphaz by protein kinase C inhibited the GAP activity of RGSZ1 and other RGS proteins, providing a mechanism for potentiation of Gz signaling by protein kinase C.
Wang
J
J
Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, Texas 75235-9041, USA.
Ducret
A
A
Tu
Y
Y
Kozasa
T
T
Aebersold
R
R
Ross
E M
EM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000378:metabolism*; D002417:Cattle; D003001:Cloning, Molecular; D003902:Detergents / Q000494:pharmacology; D004789:Enzyme Activation / Q000502:physiology; D044385:GTP-Binding Protein alpha Subunits; D019204:GTP-Binding Proteins / Q000378:metabolism*; D020690:GTPase-Activating Proteins; D020962:Heterotrimeric GTP-Binding Proteins; D006801:Humans; D008081:Liposomes / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D020710:RGS Proteins; D012333:RNA, Messenger / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D003902:Detergents; C099387:GNAZ protein, human; D044385:GTP-Binding Protein alpha Subunits; D020690:GTPase-Activating Proteins; D008081:Liposomes; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010750:Phosphoproteins; D011506:Proteins; D020710:RGS Proteins; C080831:RGS1 protein, human; C115120:RGS20 protein, human; D012333:RNA, Messenger; C097187:regulator of G-protein signalling 19; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D020962:Heterotrimeric GTP-Binding Proteins