Light-dependent phosphorylation of the carboxy tail of mouse melanopsin
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Melanopsin-based phototransduction is involved in non-image forming light responses including circadian entrainment, pupil constriction, suppression of pineal melatonin synthesis, and direct photic regulation of sleep in vertebrates. Given that the functions of melanopsin involve the measurement and summation of total environmental luminance, there would appear to be no need for the rapid deactivation typical of other G-protein coupled receptors. In this study, however, we demonstrate that heterologously expressed mouse melanopsin is phosphorylated in a light-dependent manner, and that this phosphorylation is involved in regulating the rate of G-protein activation and the lifetime of melanopsin’s active state. Furthermore, we provide evidence for light-dependent phosphorylation of melanopsin in the mouse retina using an in situ proximity ligation assay. Finally, we demonstrate that melanopsin preferentially interacts with the GRK2/3 family of G-protein coupled receptor kinases through co-immunoprecipitation assays. Based on the complement of G-protein receptor kinases present in the melanopsin-expressing retinal ganglion cells, GRK2 emerges as the best candidate for melanopsin’s cognate GRK.
KeywordsMelanopsin Phosphorylation In situ proximity assay
G-protein coupled receptor
G-protein coupled receptor kinase
Proximity-dependent ligation assay
Intrinsically photosensitive retinal ganglion cell
This study was supported by Grants from the National Science Foundation to P.R.R (IOS0721608), and from the National Eye Institute to P. R. R. (R01EY019053). J. B. was supported by an NIH training grant (T32 GM066706). We would like to thank R. Crouch for the gift of 11-cis-retinal and D. Ujla for constructing the phospho-null melanopsin mutant.
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