The p67 mRNA level and p67 requirement in protein synthesis were studied using an animal cell (KRC-7, rat tumor hepatoma cell) in culture. p67 mRNA was present in confluent cells but disappeared almost completely from serum-starved cells. However, when PMA was added to the serum-starved cells, p67 mRNA appeared in increasing quantities. Several-fold molar excess of p67 mRNA over that present in confluent cells was detected within 2 h of PMA addition and this level remained the same during the 4 h of the experiment. p67 requirement in protein synthesis was studied using a p67 antisense DNA construct under a metallothionein gene promoter. Expression of this antisense DNA in the presence of zinc in PMA-induced serum-starved cells completely inhibited induced appearance of p67 mRNA and subsequent protein synthesis. These results suggest that p67 is regulated at the mRNA level and also that this protein factor is essential for protein synthesis.