RGS17/RGSZ2, a novel regulator of Gi/o, Gz, and Gq signaling.
To identify novel regulators of Galpha(o), the most abundant G-protein in brain, we used yeast two-hybrid screening with constitutively active Galpha(o) as bait and identified a new regulator of G-protein signaling (RGS) protein, RGS17 (RGSZ2), as a novel human member of the RZ (or A) subfamily of RGS proteins. RGS17 contains an amino-terminal cysteine-rich motif and a carboxyl-terminal RGS domain with highest homology to hRGSZ1- and hRGS-Galpha-interacting protein. RGS17 RNA was strongly expressed as multiple species in cerebellum and other brain regions. The interactions between hRGS17 and active forms of Galpha(i1-3), Galpha(o), Galpha(z), or Galpha(q) but not Galpha(s) were detected by yeast two-hybrid assay, in vitro pull-down assay, and co-immunoprecipitation studies. Recombinant RGS17 acted as a GTPase-activating protein (GAP) on free Galpha(i2) and Galpha(o) under pre-steady-state conditions, and on M2-muscarinic receptor-activated Galpha(i1), Galpha(i2), Galpha(i3), Galpha(z), and Galpha(o) in steady-state GTPase assays in vitro. Unlike RGSZ1, which is highly selective for G(z), RGS17 exhibited limited selectivity for G(o) among G(i)/G(o) proteins. All RZ family members reduced dopamine-D2/Galpha(i)-mediated inhibition of cAMP formation and abolished thyrotropin-releasing hormone receptor/Galpha(q)-mediated calcium mobilization. RGS17 is a new RZ member that preferentially inhibits receptor signaling via G(i/o), G(z), and G(q) over G(s) to enhance cAMP-dependent signaling and inhibit calcium signaling. Differences observed between in vitro GAP assays and whole-cell signaling suggest additional determinants of the G-protein specificity of RGS GAP effects that could include receptors and effectors.
Mao
Helen
H
Department of Neuroscience, Ottawa Health Research Institute, University of Ottawa, Ottawa, Ontario K1H 8M5, Canada.
Zhao
Qingshi
Q
Daigle
Mireille
M
Ghahremani
Mohammad H
MH
Chidiac
Peter
P
Albert
Paul R
PR
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D003545:Cysteine / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D020558:GTP Phosphohydrolases / Q000378:metabolism; D044385:GTP-Binding Protein alpha Subunits; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D007313:Insecta; D008969:Molecular Sequence Data; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry; D051381:Rats; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D020798:Two-Hybrid System Techniques
C099387:GNAZ protein, human; D044385:GTP-Binding Protein alpha Subunits; C490963:Gnaz protein, rat; D012313:RNA; D005982:Glutathione Transferase; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D003545:Cysteine; D002118:Calcium