Light- and guanosine 5'-3-O-(thio)triphosphate-sensitive localization of a G protein and its effector on detergent-resistant membrane rafts in rod photoreceptor outer segments.
Detergent-resistant membrane microdomains in the plasma membrane, known as lipid rafts, have been implicated in various cellular processes. We report here that a low-density Triton X-100-insoluble membrane (detergent-resistant membrane; DRM) fraction is present in bovine rod photoreceptor outer segments (ROS). In dark-adapted ROS, transducin and most of cGMP-phosphodiesterase (PDE) were detergent-soluble. When ROS membranes were exposed to light, however, a large portion of transducin localized in the DRM fraction. Furthermore, on addition of guanosine 5'-3-O-(thio)triphosphate (GTPgammaS) to light-bleached ROS, transducin became detergent-soluble again. PDE was not recruited to the DRM fraction after light stimulus alone, but simultaneous stimulation by light and GTPgammaS induced a massive translocation of all PDE subunits to the DRM. A cholesterol-removing reagent, methyl-beta-cyclodextrin, selectively but partially solubilized PDE from the DRM, suggesting that cholesterol contributes, at least in part, to the association of PDE with the DRM. By contrast, transducin was not extracted by the depletion of cholesterol. These data suggest that transducin and PDE are likely to perform their functions in phototransduction by changing their localization between two distinct lipid phases, rafts and surrounding fluid membrane, on disc membranes in an activation-dependent manner.
Seno
K
K
Graduate School of Science and Technology, Department of Biology, Faculty of Science, Kobe University, Nada, Kobe 657, Japan.
Kishimoto
M
M
Abe
M
M
Higuchi
Y
Y
Mieda
M
M
Owada
Y
Y
Yoshiyama
W
W
Liu
H
H
Hayashi
F
F
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D002417:Cattle; D002784:Cholesterol / Q000302:isolation & purification / Q000378:metabolism; D003505:Cyclodextrins; D003624:Darkness; D003902:Detergents; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D008027:Light; D021962:Membrane Microdomains / Q000187:drug effects / Q000502:physiology* / Q000528:radiation effects; D021381:Protein Transport; D012374:Rod Cell Outer Segment / Q000502:physiology*; D015293:Transducin / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D047392:beta-Cyclodextrins
D003505:Cyclodextrins; D003902:Detergents; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D002784:Cholesterol; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D015293:Transducin