G1 CYCLIN-DEPENDENT KINASES ARE SUFFICIENT TO INITIATE DNA-SYNTHESIS IN QUIESCENT HUMAN FIBROBLASTS

Mammalian fibroblasts require mitogens in order to exit from GO (quies cence) and progress through the G1 phase of the cell cycle, although o nce they pass the restriction point late in G1 they can enter S phase and complete the cell cycle without mitogens [1]. Mitogenic signals ar e integrated through the GTPase Ras, which regulates the levels of cyc lin D1 [2-5], a component of the cell cycle machinery that operates du ring G1 phase by activating cyclin-dependent kinase 4 (Cdk4). The accu mulation of active cyclin E-Cdk2 complexes also requires pas [6]. Thes e two G1 cyclin-Cdk complexes act on a family of E2F-associated transc riptional repressors typified by the retinoblastoma protein (Rb) to br ing about a transcriptional program that promotes passage through S ph ase [7-9], but can also activate DNA replication independently of Rb-E 2F [10-12]. Although G1 cyclin-Cdk complexes are required for S-phase entry and can shorten G1 phase when overexpressed [13-15], it is not k nown whether they are sufficient for this transition. Here, we report that serum starved (GO) diploid human fibroblasts initiate DNA synthes is upon microinjection of active G1 cyclin-Cdk complexes, but not upon microinjection of an S phase cyclin-Cdk complex. These data indicate that G1 Cdk activation is rate-limiting for S-phase entry, and that Cd k activation is likely to be the primary function of growth factor sig nalling pathways that lead to DNA synthesis.