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This paper explores the transitional states that bridge the gap between nuclear quiescence and mitogenesis. It presents evidence for the formation of a committed but prereplicative state. Quiescent murine Swiss 3T3 cells were exposed to an external mitogenic stimulant (epidermal growth factor or excess serum) and simultaneously to a synchronizer which inhibits entry into the S phase. Thus, the cells were stimulated to synthesize DNA, but the normal replicative response to this stimulus was blocked. The block to DNA replication was removed at varying times after removal of the stimulant. Experiments were done to monitor the decay of commitment to DNA synthesis after removal of the external stimulant. This decay turned out to be a first-order process. The half-life (time required for loss of the commitment to DNA synthesis in half of the initially sensitized cells) was found to be approximately 5 hr. The same result was found whether total DNA synthesis or individually replicating cells were measured and was independent of the type of external growth stimulant or blocker used. These results point to the existence, on the mitogenic pathway, of a committed but prereplicative state. The committed state appears to represent a unit or global property of the whole cell rather than, for example, a critical concentration of some active inducer molecule because the latter would display multi-hit decay kinetics rather than the single-step lability actually observed.

Initiation of nuclear DNA replication: evidence for formation of committed prereplicative cellular state.