Extracellular adenosine 3',5'-cyclic monophosphate (cAMP) is required for cell-type-specific gene expression in developing Dictyostelium discoideum. We have developed a microassay for the expression of these genes, using antibodies directed against their protein products. To characterize the transduction mechanism, we have used in this assay cAMP analogues that preferentially activate either the cellsurface cAMP receptor or the internal cAMP-dependent pro- The mechanism whereby the extracellular cAMP induces cell-type-specific gene expression is unknown. One possible mechanism is that cAMP binds to a cell-surface receptor, which then mediates the gene induction. A cAMP-dependent protein kinase has been shown to exist in Dictyostelium (18)(19)(20)(21)(22). A second possible mechanism would then be that cAMP enters the cells and directly activates this kinase. A third possibility would involve a yet unidentified cAMPbinding protein.As an approach to these questions, we have used cAMP analogues that preferentially activate either the cell-surface cAMP receptor or cAMP-dependent protein kinase in vitro. We have established a microassay by using antibodies made against regions of the protein products of several of these cAMP-regulated cell-type-specific genes. This assay allows us to examine the ability of the analogues to activate cell-type-specific gene expression in low-density cultures measured by an immunofluorescence assay and to examine questions concerning the differential expression of these genes. We find that cAMP analogues that activate the cell-surface receptor and have very limited ability to activate the cAMP-dependent protein kinase can induce cell-typespecific gene expression, while other analogues that activate cAMP-dependent protein kinase but not the cell-surface receptor are unable to cause cell-type-specific gene expression. These low-density cultures also allow the examination of the levels of cAMP necessary for cell-type-specific gene induction.

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The phosphoproteins of Dictyostelium discoideum were compared at different stages of development of polyacrylamide gel electrophoresis. Certain phosphoproteins of vegetative amoebae were conserved while others appeared and disappeared during development. Four major phosphoproteins with apparent subunit molecular weights of 50,000, 47,000, 38,000, and 34,000 disappeared precociously in response to exogenous of cAMP. Two membranal phosphoproteins, with apparent subunit molecular weights of 80,000 and 81,000, appeared precociously in response to added cAMP. One of these phosphoproteins, molecular weight 80,000, has been identified tentatively as the "contact site A"glycoprotein. Another membranal protein, with apparent subunit molecular weight of 42,000, unaffected in its appearance by cAMP, has been identified tentatively as phosphoactin.