AUTOCRINE REGULATION OF CELL-CYCLE PROGRESSION IN NORMAL HUMAN KERATINOCYTES

Removal of competence factors insulin and pituitary extract from the c ulture medium, concomitant with the addition of picomolar concentratio ns of the late-G(1) inhibitor transforming growth factor-beta, effecti vely arrested cell cycle progression of normal human keratinocytes pri or to their entry into the DNA synthesis phase; arrest continued for a minimum of 36 h following removal of unbound inhibitor and subsequent addition of factor-deficient medium. To demonstrate the reversibility of transforming growth factor-beta-induced arrest, two dissimilar cel l populations were recruited to synthesize DNA in a predictable and re producible manner; whereas the reinstatement of omitted competence fac tors induced noncycling cells to begin synthesizing DNA within 24 h, a ddition of keratinocyte-conditioned medium prompted an immediate progr ession of late-G(1) cells into S phase. Studies to determine the exten t that autocrine signaling regulates cell cycle progression revealed t hat nontransformed keratinocytes produce an endogenous factor required for DNA replication and that production of this progression factor re quired competence factors insulin and pituitary extract. Keratinocyte progression factor recruited late-G(1) cells into S phase within 1-2 h , reversed transforming growth factor-beta-induced arrest in the prese nce of bound inhibitor, and elicited a calcium mobilization response c onsistent with receptor-mediated signaling. Hence, these studies demon strate that G(1) progression of nontransformed keratinocytes into S ph ase requires an endogenous progression factor and suggest that this fa ctor may direct G(1) progression by modulating the activity of a calci um-dependent kinase.