Regulation of Arrestin Translocation by Ca2+ and Myosin III in Drosophila Photoreceptors

Upon illumination several phototransduction proteins translocate between cell body and photosensory compartments. In Drosophila photoreceptors arrestin (Arr2) translocates from cell body to the microvillar rhabdomere down a diffusion gradient created by binding of Arr2 to photo-isomerized metarhodopsin. Translocation is profoundly slowed in mutants of key phototransduction proteins including phospholipase C (PLC) and the Ca2+-permeable transient receptor potential channel (TRP), but how the phototransduction cascade accelerates Arr2 translocation is unknown. Using real-time fluorescent imaging of Arr2-green fluorescent protein translocation in dissociated ommatidia, we show that translocation is profoundly slowed in Ca2+-free solutions. Conversely, in a blind PLC mutant with similar to 100-fold slower translocation, rapid translocation was rescued by the Ca2+ ionophore, ionomycin. In mutants lacking NINAC (calmodulin [CaM] binding myosin III) in the cell body, translocation remained rapid even in Ca2+-free solutions. Immunolabeling revealed that Arr2 in the cell body colocalized with NINAC in the dark. In intact eyes, the impaired translocation found in trp mutants was rescued in ninaC; trp double mutants. Nevertheless, translocation following prolonged dark adaptation was significantly slower in ninaC mutants, than in wild type: a difference that was reflected in the slow decay of the electroretinogram. The results suggest that cytosolic NINAC is a Ca2+-dependent binding target for Arr2, which protects Arr2 from immobilization by a second potential sink that sequesters and releases arrestin on a much slower timescale. We propose that rapid Ca2+/CaM-dependent release of Arr2 from NINAC upon Ca2+ influx accounts for the acceleration of translocation by phototransduction.