cAMP activates adenylate and guanylate cyclase of Dictyostelium discoideum cells by binding to different classes of cell-surface receptors. A study with extracellular Ca2+.
cAMP induces a transient increase of cAMP and cGMP levels in Dictyostelium discoideum cells. Fast binding experiments reveal three types of cAMP-binding site (S, H and L), which have different off-rates (t0.5, 0.7-15 s) and different affinities (Kd, 15-450 nM). A time- and cAMP-concentration-dependent transition of H- to L-sites occurs during the binding reaction (Van Haastert, P.J.M. and De Wit, R.J.W. (1984) J. Biol. Chem. 13321-13328). Extracellular Ca2+ had multiple effects on cAMP-binding sites. (i) The number of H + L-sites increased 2.5-fold, while the number of S-sites was not strongly affected. (ii) The Kd of the S-sites was reduced from 16 nM to 5 nM (iii) The conversion of H-sites to L-sites was inhibited up to 80%. The kinetics of the cAMP-induced cAMP accumulation was not strongly altered by Ca2+, but the amount of cAMP produced was inhibited up to 80%. The kinetics of the cAMP-induced cGMP accumulation was strongly altered; maximal levels were obtained sooner, and the Ka was reduced from 15 to 3.5 nM cAMP. Ca2+, Mg2+ and Mn2+ increased the number of binding sites, all with EC50 = 0.5 mM. The S-sites and the cGMP response were modified by equal Ca2+ concentrations and by higher concentrations of Mg2+ and Mn2+ (EC50 are respectively 0.4 mM, 2.5 mM and about 25 mM). The conversion of H- to L-sites and the cAMP response were specifically inhibited by Ca2+ with EC50 = 20 microM. It is concluded that cAMP activates guanylate cyclase through the S-sites; adenylate cyclase is activated by the H + L-sites, in which the appearance of the L-sites during the binding reaction represents the coupling of occupied surface cAMP receptors to adenylate cyclase.
van Haastert
P J
PJ
D000262:Adenylyl Cyclases / Q000378:metabolism*; D002118:Calcium / Q000494:pharmacology*; D000242:Cyclic AMP / Q000378:metabolism / Q000494:pharmacology*; D006152:Cyclic GMP / Q000494:pharmacology; D004023:Dictyostelium / Q000201:enzymology / Q000378:metabolism*; D004789:Enzyme Activation; D006162:Guanylate Cyclase / Q000378:metabolism*; D007700:Kinetics; D011953:Receptors, Cyclic AMP / Q000378:metabolism*
D011953:Receptors, Cyclic AMP; D000242:Cyclic AMP; D000262:Adenylyl Cyclases; D006162:Guanylate Cyclase; D006152:Cyclic GMP; D002118:Calcium