Specific photoaffinity labeling of the cAMP surface receptor in Dictyostelium discoideum.
The recent observation that ammonium sulfate stabilizes cell-surface [3H]cyclic AMP binding in Dictyostelium discoideum (Van Haastert, P., and Kien, E. (1983) J. Biol. Chem. 258, 9636-9642) led us to attempt to identify the surface cAMP receptor by photoaffinity labeling with 8-azido-[32P]cAMP using this stabilization technique. 8-azido-[32P]cAMP specifically labeled a polypeptide which migrates as a closely spaced doublet (Mr = 40,000 to 43,000) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Greater than 60% of the labeled polypeptide was found associated with membranes. This protein was distinguished from the cytosolic regulatory subunit of the cAMP-dependent protein kinase (Mr = 41,000) by differences in developmental regulation, specificity, and subcellular localization. No kinase regulatory subunit was detected in membranes by western blot analysis. Our preliminary observations show that labeling of this doublet correlates closely with cAMP-binding activity, suggesting that it is the surface receptor which mediates chemotaxis and cAMP signaling.
D001386:Azides / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000378:metabolism; D004023:Dictyostelium / Q000378:metabolism*; D046911:Macromolecular Substances; D010761:Phosphorus Radioisotopes; D011494:Protein Kinases / Q000378:metabolism; D011953:Receptors, Cyclic AMP / Q000378:metabolism*; D014316:Tritium
D001386:Azides; D046911:Macromolecular Substances; D010761:Phosphorus Radioisotopes; D011953:Receptors, Cyclic AMP; D014316:Tritium; C017042:8-azidoadenosine-3',5'-monophosphate; D000242:Cyclic AMP; D011494:Protein Kinases