Transient Superinducibility of Cytochrome P450c (CYP1A1) mRNA and Transcription

Abstract

Simultaneous treatment of the rat hepatocyte-derived cell line LCS7 with cycloheximide and polycyclic aromatic compounds increased CYP1A1 (cytochrome P4S0c) gene transcription rate four- to sixfold and mRNA levels 20-fold relative to the levels in cells treated with inducers alone. When cycloheximide was added up to 1 hr after inducer, a similar degree of superinduction occurred. However, if cycloheximide was added at 1.5 hr or later, superinduction did not occur, even though an increased transcription rate continued at these times in cells treated with inducers alone. Thus, treatment with cycloheximide revealed two phases to the response of the CYP1A1 gene to inducer. During the early phase, inhibition of protein synthesis could amplify the effect of inducer. During the later phase, transcription rate and CYP1A1 mRNA levels remained elevated due to inducer treatment, but could not be further elevated by inhibiting protein synthesis. The superinduction of CYP1A1 mRNA was also examined in primary hepatocyte cultures and in explant cultures of three tissues. These varied substantially in their superinduction response. All of these exhibited elevated levels of CYP1A1 mRNA following simultaneous treatment with inducer and cycloheximide; transcription rate was superinduced three- to fourfold in primary hepatocytes, and CYP1A1 mRNA levels were superinduced 20- to 25-fold in both kidney and lung explants. However, the delayed addition of cycloheximide had varying effects in different culture systems. In primary hepatocytes, addition of cycloheximide as long as 4 hr after addition of inducer resulted in superinduction equal to that which occurred when these agents were added together. In contrast, adding cycloheximide only 2.5 hr after adding inducer resulted in undetectable superinduction in kidney explants and diminished superinduction 10% in lung explants. Although the time course of cycloheximide responsiveness following inducer treatment has been studied in detail only in LCS7 cells, it appears that the length of the cycloheximide-responsive phase is different in different cell culture systems in in explants from different tissues. This may be related to the previously reported tissue-specific differences in CYP1A1 gene expression.