We have identified genes that are expressed preferentially in either prestalk or prespore cells in Dictyostelium. The prestalk mRNAs are detectable at 7.5 hr prior to the completion of cell aggregation, while the prespore mRNAs are not detectable until approximately 15 hr of development. Exogenous cAMP in the absence of sustained cell contact is sufficient to induce prestalk-specific gene expression, while multicellularity is required for the induction of prespore-specific genes. A gene expressed equally in both cell types, which has the same developmental kinetics as the prestalk genes, is induced in shaking culture in the absence of either cAMP or stable cell associations. Dissociation of aggregates results in the rapid loss of prespore- and prestalk-specific mRNAs, and these can be induced to reaccumulate with the addition of cAMP. We conclude that there are substantial differences in the timing and requirements for tissue-specific gene expression in Dictyostelium.