DIFFERENTIAL REGULATION OF P27 AND CYCLIN D1 BY TGF-BETA AND EGF IN C3H 10T1 2 MOUSE FIBROBLASTS/

Previously, we found that stimulation of C3H 10T1/2 mouse fibroblasts with TGF-beta leads to the striking and rapid down-regulation of p27(K ip1) expression during G1 phase. Here, we demonstrate that TGF-beta tr eatment of C3H 10T1/2 cells does not alter the steady-state level of K ip1 message sufficiently to account for the observed down-regulation o f p27. This demonstrates that TGF-beta-induced down regulation of p27( Kip1) occurs at a post-transcriptional level, consistent with a degrad ative mechanism of p27(Kip1) down-regulation. Epidermal growth factor (EGF) does not lead to the rapid down-regulation of p27 observed follo wing treatment of cells with TGF-beta. Also in contrast with TGF-beta, EGF causes a strong upregulation of cyclin D1, while neither growth f actor affects cdk4 protein levels. These results imply that in this ce ll type TGF-beta overcomes an inhibitory threshold to cdk activation b y cyclin-dependent kinase inhibitors primarily through down-regulation of p27, while EGF overcomes this threshold predominantly through upre gulation of cyclin D1 levels. This divergence in pathways may explain why TGF-beta-induced cell cycle kinetics are slower than those of EGF in these cells, and the ability of TGF-beta to delay EGF-induced cell cycle kinetics to its own, slower kinetics. In support of this hypothe sis, TGF-beta prevents EGF-induced upregulation of cyclin D1 levels, w hile TGF-beta is still able to induce p27 down-regulation even in the presence of EGF. In contrast to the case with p27 degredation, neither TGF-beta nor EGF have an observable effect on the steady-state levels of p21 in this cell type. (C) 1996 Wiley-Liss, Inc.