Phosphorylation reactions in bovine rod outer segments studied by 32P-labelling of intact retina.
The protein phosphorylation pattern in the intact bovine retina has been investigated by labelling with 32P-phosphate under incubation conditions that preserve the electrical photoresponse of the photoreceptor cells. The phosphorylation of rod outer segment proteins was analysed after isolation of outer segments from the labelled retina. The global influence of light, Ca2+ and the phosphodiesterase inhibitor, isobutylmethylxanthine, on protein phosphorylation in rod outer segments was analysed. A 12 kDa protein is the most prominent phosphorylated species in the intact bovine retina. Its phosphorylation is increased by light and/or Ca2+. Evidence is presented that this strongly phosphorylated protein is not located in the outer segment, and we suggest that it may be a synaptic protein. Retinal rod outer segment membrane proteins with apparent molecular weights of 245, 226, 125, 110, 50, 46, 38 and 20 all show light-stimulated phosphorylation. Lowering the extracellular Ca2+ levels results in a decrease of the phosphorylation level of some of these proteins, viz. at 125, 50, 38 and probably at 20 kDa. Such proteins, whose phosphorylation level is influenced both by light and by elevated Ca2+, are candidates for mediators of phototransduction. The phosphorylated species at 245, 226, 110, 50 and 20 kDa are enriched in rod outer segment plasma membrane preparations. These protein species could participate in the light-regulated modulation of the Na+-conductance of the plasma membrane.
K M
KM
Kamps
E A
EA
Dratz
F J
FJ
Daemen
W J
WJ
de Grip
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002417:Cattle; D002499:Centrifugation, Density Gradient; D006152:Cyclic GMP / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004594:Electrophysiology; D008027:Light; D008565:Membrane Proteins / Q000378:metabolism; D008970:Molecular Weight; D010766:Phosphorylation; D010786:Photoreceptor Cells / Q000378:metabolism*; D012160:Retina / Q000378:metabolism*; D012374:Rod Cell Outer Segment / Q000378:metabolism*
D008565:Membrane Proteins; D006152:Cyclic GMP; D002118:Calcium; D015056:1-Methyl-3-isobutylxanthine