Ligand-induced Modification of a Surface cAMP Receptor of Dictyostelium discoideum Does Not Require Its Occupancy

In Dictyostelium discoideum amoebae, cAMP-induced phosphorylation of the surface cAMP receptor is associated with a discrete transition in its electrophoretic mobility. The native and modified forms of the receptor are designated R and D (Mr = 40,000 and 43,000). The relationship of the number of receptors which are modified as a function of the receptors which bind cAMP was investigated. Modification was assessed by determining the amounts of R and D forms in Western blots which detect all receptors whether or not they are exposed on the surface. Cyclic AMP or the analog, adenosine 3',5’-monophosphorothioate ((Rp)-cAMPS), induced a loss of cAMP-binding activity (down-regulation), which was not accompanied by a loss of the receptor protein. About 60% of the receptors do not bind cAMP in the absence of Ca2+ and are unmasked by 10 mM Ca2+. However, the fraction of receptors which are modified in response to cAMP is equal in the absence or presence of Ca2+. (Rp)-cAMPS induces down-regulation (50%) but not modification. Addition of cAMP, following down-regulation by (Rp)-cAMPS, causes all receptors to be modified. cAMP induces both down-regulation (80%) and modification. Modification is more readily reversed than down-regulation: 30 min after removal of cAMP, receptors remain down-regulated (57%) but are found in the R form. All receptors shift to the D form when cAMP is readded to the cells. These results indicate that exposed, as well as cryptic and down-regulated receptors, are modified in response to the cAMP stimulus