AbstractAfter retinoic acid treatment, a large percentage of cells of the human embryonal carcinoma cell line NT2/D1 differentiate into neuronal cells. We demonstrate here that the differentiated cells, but not the undifferentiated cells, contain high levels of neurofilament mRNA. We have also measured mRNA, protein, and activity levels of two kinases, cAMP‐dependent protein kinase (PKA) and protein kinase C (PKC), in order to explore the role of protein kinases in the establishment of the differentiated state. RNA levels for the catalytic (Cα and Cβ) subunits of PKA increased after differentiation. Total PKA activity levels increased 7‐fold in the differentiated cells. Parallel with this, a rise in the level of catalytic subunit protein occurred. A 12‐fold induction of Type 2 (β) PKC mRNA levels was observed after neuronal differentiation. Increases in PKC activity and in Type 2 (β) and Type 3 (α) PKC protein levels also accompa nied differentiation. These changes in PKA‐ and PKC‐specific RNA levels and enzyme activity may be necessary for production and maintenance of the differentiated state