INHIBITION OF TRANSCRIPTION BLOCKS CELL-CYCLE PROGRESSION OF NIH3T3 FIBROBLASTS SPECIFICALLY IN G(1)

We have analysed the role of RNA polymerase II-dependent transcription in cell cycle progression. Time-lapse video recording and cytogenetic analysis were used to determine the sensitivity of NIH3T3 cells to th e RNA polymerase II inhibitor alpha-amanitin at different stages of th e cell cycle. Our results show that alpha-amanitin blocks cells specif ically in G1, irrespective of the concentration within the range of 3 to 30 mug/ml. This indicates that transcription in G1 is required to o vercome a restriction point located in this phase of the cell cycle. I n agreement with this conclusion is the requirement for an uninhibited protein synthesis during G1 progression. In addition, the insensitivi ty of S-phase cells to RNA polymerase II inhibition suggests that the transcription of genes thought to be normally induced during S/G2 is n ot required for the completion of an ongoing cell cycle. S/G2 progress ion was however clearly dependent on protein synthesis. This suggests that cells exposed to alpha-amanitin are able to complete their cell c ycle because sufficiently high levels of mRNA are present in S/G2 due to basal level transcription, or are left from preceding cell cycles. It is therefore unlikely that transcriptional regulation in S or G2 pl ays a crucial role in the control of cell cycle progression in NIH3T3 cells.