Ubiquitylation of the transducin βγ subunit complex: Regulation by phosducin

Abstract

G proteins (Gαβγ) are essential signaling molecules, which dissociate into Gα and Gβγ upon activation by heptahelical membrane receptors. We have identified the βγ subunit complex of the photoreceptor-specific G protein, transducin (T), as a target of the ubiquitin-proteasome pathway. Ubiquitylated species of the transducin γ-subunit (Tγ) but not the α- or β-subunits were assembled de novo in bovine photoreceptor preparations. In addition, Tγ was exclusively ubiquitylated when Tβγ was dissociated from Tα. Ubiquitylation of Tβγ on Tγ was selectively catalyzed by human ubiquitin-conjugating enzymes UbcH5 and UbcH7 and was coincident with degradation of the entire Tβγ subunit complex in vitro by a mechanism requiring ATP and the proteasome. We also show that Tβγ association with phosducin, a photoreceptor-specific protein of unknown physiological function, blocks Tβγ ubiquitylation and subsequent degradation. Phosphorylation of phosducin by Ca2+/calmodulin-dependent protein kinase II, which inhibits phosducin-Tβγ complex formation, completely restored Tβγ ubiquitylation and degradation. We conclude that Tβγ is a substrate of the ubiquitin-proteasome pathway and suggest that phosducin serves to protect Tβγ following the light-dependent dissociation of Tαβγ.