Protein kinase C-mediated phosphorylation of retinal rod outer segment membrane proteins

In the present study we demonstrate that TD-α undergoes phosphorylation by PKC when present in its native form in intact ROS membranes. This phosphorylation is inhibited by GTP-γ-S which activates TD, suggesting that it is only the inactive conformation of TD-α that serves as a substrate for PKC. Indeed, both vanadate and AlF₄, that confer an active conformation on TD-α-GDP, inhibit PKC-mediated phosphorylation of purified TD-α-GDP. We demonstrate that the purified β subunit of TD also serves as an in vitro substrate for PKC. Moreover, following their phosphorylation, both TD-α and β from high affinity complexes with PKC. This is evident from the findings that PKC coprecipitates with both the α and β subunits of TD when the latter are immunoprecipitated by their respective antibodies. PKC phosphorylates additional ROS proteins of 36, 48 and 92 kDa, tentatively identified as rhodopsin, arrestin and the cGMP-phosphodiesterase. Taken together our results strongly suggest that phosphorylation of TD is of physiological relevance and that through phosphorylation of endogenous ROS proteins, PKC could play a key role in regulating phototransduction.