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J.J. Peterson, O.L. Moritz, B. Tam, D.S. Papermaster, W.C. Smith; Involvement of Cytoskeletal Elements in the Complementary Migration of Arrestin and Transducin in Xenopus Photoreceptors . Invest. Ophthalmol. Vis. Sci. 2003;44(13):4262.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: To study the mechanisms governing the migrations of arrestin and transducin in Xenopus rod photoreceptors in response to light. Methods: Immunohistochemical techniques employing chaotropic agents and prolonged histochemical exposure were used that faithfully reproduced the results previously obtained using an arrestin-GFP fusion in transgenic Xenopus tadpoles. Dark-adapted tadpoles were incubated in Ringers containing 0.2% DMSO, with or without 25 µM cytochalasin D to inhibit actin polymerization for 6 h. Tadpoles were then either 1) dark-adapted (DA) for an additional 30 min, 2) light-adapted (LA) for 45 min, or 3) LA for 4 h and fixed. Cryosections were treated with 0.1% NaBH4, 1% Triton X-100, and 6 M guanidinium-HCl. Sections were stained with arrestin and transducin G-α antibodies and visualized using fluorescent confocal microscopy. Results: In wild-type tadpoles, immunocytochemically-stained arrestin was localized as previously reported using transgenic tadpoles expressing arrestin-GFP; i.e., arrestin localization shifts from the inner segment (IS) to the outer segment (OS) after 45 min of LA but returns to the IS and axoneme in response to extended LA. We demonstrate for the first time that transducin movement is fully reciprocal to that of arrestin; i.e. transducin resides in the OS in the dark, migrates to the IS on brief light exposure and then relocates to the OS after prolonged exposure. Transducin in untreated tadpoles was localized exclusively in the OS in the dark in a hatched pattern corresponding to the lateral alignment of disc membranes. After 45 min of LA, transducin appeared to spread and intensify throughout the OS and was detected throughout the IS, including the synapse. After extended LA, transducin was again localized exclusively in the OS. Unlike arrestin, transducin localization was sensitive to cytochalasin D, which caused transducin distribution to broaden and intensify in the OS in the dark, and inhibited complete relocalization to the OS in response to extended light exposure. Conclusions: Migrations of arrestin and transducin are complementary. Even during extended LA, transducin and arrestin migrations are polarized in Xenopus tadpoles. Treatment of tadpoles with cytochalasin D disrupted the light-activated movement of transducin, but not arrestin. These observations suggest that light induced transducin movement involves microfilaments but arrestin migration does not.
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