Regulatory sequences of a sea urchin cytoskeletal actin gene (CyIIIa) were ligated to the bacterial gene coding for chloramphenicol acetyltransferase (CAT; acetyl-CoA:chloramphenicol O3-acetyltransferase, EC 2.3.1.28) and the construct was injected into unfertilized sea urchin eggs. CAT activity is detected at the early blastula stage, when transcripts of the endogenous CyIIIa gene normally appear. Our measurements show that during activation the amount of CAT enzyme increases at least 100-fold; that there are present in late blastula stage embryos about 5 X 10(5) molecules of CAT mRNA (i.e., approximately 6 times the number of endogenous CyIIIa mRNAs); and that within the range studied the amount of CAT enzyme produced is independent of the number of CyIIIa-CAT genes incorporated per embryo, probably because the genes are present in excess of factors required for their activation. Activation of the CyIIIa-CAT construct is seriously inhibited, or abolished, by successive deletions of upstream CyIIIa sequences.