Activation-dependent hindrance of photoreceptor G protein diffusion by lipid microdomains.
The dynamics of G protein-mediated signal transduction depend on the two-dimensional diffusion of membrane-bound G proteins and receptors, which has been suggested to be rate-limiting for vertebrate phototransduction, a highly amplified G protein-coupled signaling pathway. Using fluorescence recovery after photobleaching (FRAP), we measured the diffusion of the G protein transducin alpha-subunit (Galpha(t)) and the G protein-coupled receptor rhodopsin on disk membranes of living rod photoreceptors from transgenic Xenopus laevis. Treatment with either methyl-beta-cyclodextrin or filipin III to disrupt cholesterol-containing lipid microdomains dramatically accelerated diffusion of Galpha(t) in its GTP-bound state and of the rhodopsin-Galphabetagamma(t) complex but not of rhodopsin or inactive GDP-bound Galphabetagamma. These results imply an activity-dependent sequestration of G proteins into cholesterol-dependent lipid microdomains, which limits diffusion and exclude the majority of free rhodopsin and the free G protein heterotrimer. Our data offer a novel demonstration of lipid microdomains in the internal membranes of living sensory neurons.
Wang
Qiong
Q
Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, Texas 77030, USA.
Zhang
Xue
X
Zhang
Li
L
He
Feng
F
Zhang
Guowei
G
Jamrich
Milan
M
Wensel
Theodore G
TG
D000818:Animals; D002784:Cholesterol / Q000378:metabolism; D003720:Densitometry; D019204:GTP-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000737:chemistry; D007313:Insecta; D008055:Lipids / Q000737:chemistry*; D021962:Membrane Microdomains / Q000737:chemistry*; D008856:Microscopy, Fluorescence; D009474:Neurons / Q000378:metabolism; D010786:Photoreceptor Cells / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012243:Rhodopsin / Q000737:chemistry; D015398:Signal Transduction; D014982:Xenopus laevis
D008055:Lipids; D006160:Guanosine Triphosphate; D012243:Rhodopsin; D002784:Cholesterol; D019204:GTP-Binding Proteins