Developmentally regulated compartmentalization of adenylate cyclase in Dictyostelium discoideum.
Adenylate cyclase of aggregation phase Dictyostelium discoideum is activated by extracellular adenosine 3', 5'-cyclic monophosphate (cAMP), and the cAMP synthesized is secreted. The distribution of the enzyme was determined in sucrose gradients loaded with whole cell lysates. Cell lysates prepared after 4.5 hr of starvation revealed membranes containing adenylate cyclase at 44% and 33% sucrose. The activity of the latter peak was detected in the presence of the detergent (CHAPS), 3-(3-cholamidopropyl) dimethylammonio-3-propanesulfonate, which inhibited the activity of the former to some extent. Adenylate cyclase activity of the 2 peaks differed with respect to solubility in CHAPS and their kinetics. The 44% sucrose region of the gradient contained the bulk of the plasma membranes, as judged by a cell surface glycoprotein marker (contact site A). The 33% peak is composed of small vesicular structures, as determined by electron microscopy. The distribution of adenylate cyclase activity detected in sucrose gradients shifted from the 33% to the 44% sucrose peak during development. In addition, the 44% peak became increasingly resistant to the inhibitory effect of CHAPS. Both changes were accelerated by extracellular cAMP, but only the latter was abolished when the production of endogenous cAMP was inhibited by caffeine. Pulsing cells with cAMP overcame the inhibitory effect of caffeine.
Max Planck Institute of Biochemistry, Martinsried, Federal Republic of Germany.
J
J
Hagmann
D000262:Adenylyl Cyclases / Q000302:isolation & purification / Q000378:metabolism*; D002110:Caffeine / Q000494:pharmacology; D004023:Dictyostelium / Q000201:enzymology / Q000254:growth & development* / Q000648:ultrastructure; D007700:Kinetics; D008854:Microscopy, Electron
D002110:Caffeine; D000262:Adenylyl Cyclases