The E2F1 transcription factor regulates transit of cells through the S phase checkpoint, dependent on its association with cyclin A/cdk2. Expression in cells of a mutant E2F1 lacking the cyclin A/cdk2 binding domain leads to partial arrest of cells at the S phase checkpoint. When subconfluent growing cells expressing this mutant E2F1 are analyzed in detail, it is shown here that they display a significantly reduced incorporation of 3H-thymidine into the DNA of each S phase cell, compared to control cells or to cells overexpressing full-length E2F1. Further, when cells are blocked at the G1/S phase border and released, there is a clear reduction in the amount of 3H-thymidine incorporated into the DNA of S phase cells by 1.5 hours post release. Considering a normal 6 hour S phase duration, the results show that the S phase checkpoint mediated by E2F1 is not a late S phase event but an early one.
Copyright 1999 Academic Press.