In situ assay of light-stimulated G protein activity in Drosophila photoreceptor G protein beta mutants.
An in situ 35S-labeled guanosine 5'-3-O-(thio)triphosphate (GTP gamma S) binding procedure was developed to assay light-stimulated G protein activity in Drosophila compound eyes. We found that Drosophila with mutations in G beta e, an abundant photoreceptor-specific G protein beta subunit essential for photoexcitation, are defective in light-stimulated [35S]GTP gamma S binding. We confirmed that G beta e interacts with a GTP-binding protein by demonstrating that immunoprecipitation of G beta e is sensitive to GTP gamma S. These results suggest that G beta e functions as the beta subunit of a heterotrimeric G protein that couples photoactivation of rhodopsin to downstream components in the Drosophila phototransduction cascade.
Yarfitz
S L
SL
Department of Biochemistry, University of Washington, Seattle 98195.
Running Deer
J L
JL
Froelick
G
G
Colley
N J
NJ
Hurley
J B
JB
D000818:Animals; D001345:Autoradiography; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D004330:Drosophila / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D019204:GTP-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism* / Q000528:radiation effects; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism*; D015151:Immunoblotting; D008027:Light; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017956:Photoreceptor Cells, Invertebrate / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D013462:Sulfur Radioisotopes
D017931:DNA Primers; D011994:Recombinant Proteins; D013462:Sulfur Radioisotopes; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D019204:GTP-Binding Proteins