ROLE OF CDC25-C PHOSPHATASE IN THE IMMEDIATE G2 DELAY INDUCED BY THE EXOGENOUS FACTORS EPIDERMAL GROWTH-FACTOR AND PHORBOLESTER

Studies on the link between cellular signalling and cell cycle control at the G2 checkpoint have shown that, in HeLa cells, epidermal growth factor (EGF) and the phorbol ester 12-O-tetradecanoylphorbol-13-aceta te (TPA) rapidly inhibit the G2-M transition by preventing the key com ponent of mitosis-promoting factor (MPF), p34(cdc2), from expressing p rotein kinase activity. The kinase activity of active MPF is not inhib ited; rather, the conversion of pre-MPF to MPF, i.e., the activating d ephosphorylation of p34(cdc2), at tyrosine is rapidly blocked (Barth a nd Kinzel, 1994, Exp. Cell Res. 212:383-388; 1995, J. Cell. Physiol., 162:44-51). The phosphatase responsible, cdc25-C, is activated by phos phorylation in mitotic cells starting at the G2-M transition in an aut ocatalytic loop with MPF (Hoffmann et al., 1993, EMBO J. 12:53-63). We now show that, concomitant with the prevention of MPF activation, EGF and TPA induced a reduction of the activity of cdc25-C in synchronize d cultures. Furthermore, treatment of mitotic HeLa cells with TPA did not influence the kinase activity of MPF but caused a rapid decrease o f the specific enzyme activity of cdc25-C, probably due to dephosphory lation of the enzyme, as indicated by reduced binding of monoclonal MP M-2 antibody specific for phosphoepitopes in M phase. Because of its i nability to induce signalling during division, EGF failed to influence the activity of cdc25-C in mitotic cells. The scenario in cells late in G2 that are committed to enter mitosis may be as follows: In those cells where the signalling pathways responding to EGF as well as those responding to TPA are still open, cdc25-C is prevented by dephosphory lation from exceeding the threshold level of activity required to init iate the activation of and the autocatalytic feedback loop with p34 an d to enter mitosis. Therefore, cdc25-C appears to represent part of an interface between cellular signalling and cell cycle control in G2 ph ase. (C) 1996 Wiley-Liss, Inc.