Phosphorylation of proteins on tyrosine residues has been shown to govern many cellular processes, but little work has focused on the role of tyrosine phosphorylation during germination. Under optimal conditions, D. discoideum spores synchronously germinate each liberating a single amoeba. The total amount of phosphotyrosine containing proteins observed in spores was greatest during quiescence with a gradual decline during spore activation and emergence of nascent amoeba. During dormancy, tyrosine residues of actin were heavily phosphorylated, but they gradually underwent dephosphorylation upon spore activation and this process continued through emergence. Interestingly, an endogenous autoinhibitor(s), which blocks germination, induces tyrosine phosphorylation of actin. Conversely, the removal of the autoinhibitor(s) was followed by a decrease in phosphorylation. Thus, during germination of Dictyostelium spores, actin is dephosphorylated, with the level of phosphorylation regulated by the autoinhibitor(s) and/or the autoactivator. This change in actin phosphorylation appears to play a direct role since actin dephosphorylation and reorganization is a necessary prelude to germination.