KINETICS OF EXPRESSION OF INDUCIBLE BETA-GALACTOSIDASE IN MURINE FIBROBLASTS - HIGH INITIAL RATE COMPARED TO STEADY-STATE EXPRESSION

We have constructed a murine fibroblast cell line in which synthesis o f beta-galactosidase can be induced by incubation with isopropyl-beta- D-thiogalactopyranoside (IPTG). This was obtained by transfection by b oth a plasmid expressing lacI and a second plasmid expressing lacZ fro m a modified simian virus 40 (SV40) promoter containing a Iac operator . We have measured the induction kinetics as well as the basal and ind uced differential rate of synthesis of beta-galactosidase. The steady- state rate of synthesis is tenfold higher in the presence than in the absence of inducer; we calculate an average of 1200 lacZ polypeptides are synthesized per minute per cell in the induced cultures. However, immediately after induction, the rate of accumulation of beta-galactos idase is up to 50-fold higher than the basal level. Based on our measu rements of stability of beta-galactosidase, we suggest that induction may result in a subsequent down-modulation of the transcriptional acti vity from the induced gene. We hypothesize this inhibition may result from structural changes in DNA components, such as nucleosomes. (C) 19 95 Academic Press Limited.