Functional regulation of Galpha16 by protein kinase C.
Recent evidence demonstrates that the alpha subunits of some heterotrimeric GTP-binding proteins (G proteins) are subject to modification by protein kinase C (PKC). For the family of G proteins that activate the phospholipase C/inositol trisphosphate/calcium/PKC pathway, such modification could result in G protein autoregulation. To examine the potential regulation of members of the Galphaq family by PKC phosphorylation, we expressed the thyrotropin-releasing hormone (TRH) receptor in combination with Galphaq, Galpha11, Galpha14, Galpha15, or Galpha16 in Xenopus oocytes and examined the regulation of signaling by PKC activators and inhibitors. For Galpha16 and Galpha15, the two family members of hematopoietic lineage, PKC activators reduce both the magnitude and the time course of TRH-mediated responses; PKC inhibitors have the opposite effect. The PKC-mediated effects are evident in measurements of GTPase activity, suggesting that the regulation is occurring early in the signaling pathway. In vivo phosphorylation experiments demonstrate that Galpha16 is a substrate for PKC modification. By comparison, Galphaq is not phosphorylated by PKC in vivo, and oocytes expressing Galphaq are not functionally modulated by PKC. Repeated TRH stimulation of oocytes expressing Galpha16 mimics the effects of PKC activators, and this functional regulation is correlated with an increase in Galpha16 phosphorylation. A mutant Galpha16 with four consensus PKC phosphorylation sites removed is not phosphorylated in vivo, and TRH responses mediated through the mutant are not regulated by PKC. These results demonstrate that signaling involving hematopoietic G proteins is subject to PKC-mediated autoregulation, at least in part, by phosphorylation of the G protein alpha subunit.
Aragay
A M
AM
Centro de Biologia Molecular, Facultad de Ciencias, Universidad Autonoma de Madrid, Canto Blanco, 28049 Madrid, Spain.
Quick
M W
MW
D000595:Amino Acid Sequence; D000818:Animals; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D009865:Oocytes / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction; D019311:Staurosporine / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014981:Xenopus