Ubiquitylation of the transducin betagamma subunit complex. Regulation by phosducin.
G proteins (Galphabetagamma) are essential signaling molecules, which dissociate into Galpha and Gbetagamma upon activation by heptahelical membrane receptors. We have identified the betagamma subunit complex of the photoreceptor-specific G protein, transducin (T), as a target of the ubiquitin-proteasome pathway. Ubiquitylated species of the transducin gamma-subunit (Tgamma) but not the alpha- or beta-subunits were assembled de novo in bovine photoreceptor preparations. In addition, Tgamma was exclusively ubiquitylated when Tbetagamma was dissociated from Talpha. Ubiquitylation of Tbetagamma on Tgamma was selectively catalyzed by human ubiquitin-conjugating enzymes UbcH5 and UbcH7 and was coincident with degradation of the entire Tbetagamma subunit complex in vitro by a mechanism requiring ATP and the proteasome. We also show that Tbetagamma association with phosducin, a photoreceptor-specific protein of unknown physiological function, blocks Tbetagamma ubiquitylation and subsequent degradation. Phosphorylation of phosducin by Ca(2+)/calmodulin-dependent protein kinase II, which inhibits phosducin-Tbetagamma complex formation, completely restored Tbetagamma ubiquitylation and degradation. We conclude that Tbetagamma is a substrate of the ubiquitin-proteasome pathway and suggest that phosducin serves to protect Tbetagamma following the light-dependent dissociation of Talphabetagamma.
Obin
Martin
M
Laboratory for Nutrition & Vision Research, JMUSDA-HNRCA at Tufts University and Tufts Center for Vision Research, Boston, Massachusetts 02111, USA. mobin@hnrc.tufts.edu
Lee
Bruce Y
BY
Meinke
Gretchen
G
Bohm
Andrew
A
Lee
Rehwa H
RH
Gaudet
Rachelle
R
Hopp
Johnathan A
JA
Arshavsky
Vadim Y
VY
Willardson
Barry M
BM
Taylor
Allen
A
D000818:Animals; D002417:Cattle; D003600:Cytosol / Q000378:metabolism; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005136:Eye Proteins / Q000737:chemistry* / Q000378:metabolism*; D020555:GTP-Binding Protein Regulators; D010750:Phosphoproteins / Q000737:chemistry* / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D020033:Protein Isoforms; D017948:Retinal Rod Photoreceptor Cells / Q000378:metabolism; D015293:Transducin / Q000737:chemistry* / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism*
D005136:Eye Proteins; D020555:GTP-Binding Protein Regulators; D010750:Phosphoproteins; D020033:Protein Isoforms; D025801:Ubiquitin; C065631:phosducin; D015293:Transducin