It is generally accepted that the multiple, similar protein kinase C (PKC) isozymes are responsible for different specialized physiological processes, but evidence that directly assigns specific functions to specific isozymes is scarce. To test whether specific PKC isozymes are involved in myeloid differentiation, we have studied the effect of overexpression of PKC-α, -βII, -δ, -ε, -ζ and -η in 32D, a mouse myeloid progenitor cell line that does not differentiate in response to 12-O-tetradecanoylphorbol-13-acetate (TPA). No significant morphological or phenotypic changes could be observed in unstimulated cells that overexpress any of these isozymes. However, the cell lines that overexpressed PKC-α or -δ had acquired the ability to become mature macrophages 2-6 h after TPA stimulation. The overexpression of PKC-βII, -ε, -ζ, or -η, in contrast, did not permit TPA-induced differentiation. These results indicate that only these two members of the PKC gene family can participate in TPA-induced myeloid differentiation.