Scaffolding Protein INAD Regulates Deactivation of Vision by Promoting Phosphorylation of Transient Receptor Potential by Eye Protein Kinase C inDrosophila

Drosophilavisual signaling is one of the fastest G-protein-coupled transduction cascades, because effector and modulatory proteins are organized into a macromolecular complex (“transducisome”). Assembly of the complex is orchestrated by inactivation no afterpotential D (INAD), which colocalizes the transient receptor potential (TRP) Ca2+channel, phospholipase Cβ, and eye protein kinase C (eye-PKC), for more efficient signal transduction. Eye-PKC is critical for deactivation of vision. Moreover, deactivation is regulated by the interaction between INAD and TRP, because abrogation of this interaction inInaDp215results in slow deactivation similar to that ofinaCp209lacking eye-PKC. To elucidate the mechanisms whereby eye-PKC modulates deactivation, here we demonstrate that eye-PKC, via tethering to INAD, phosphorylates TRPin vitro. We reveal that Ser982of TRP is phosphorylated by eye-PKCin vitroand, importantly, in the fly eye, as shown by mass spectrometry. Furthermore, transgenic expression of modified TRP bearing an Ala substitution leads to slow deactivation of the visual response similar to that ofInaDp215. These results suggest that the INAD macromolecular complex plays an essential role in termination of the light response by promoting efficient phosphorylation at Ser982of TRP for fast deactivation of the visual signaling.