ECTOPIC EXPRESSION OF CYCLIN D1 BUT NOT CYCLIN-E INDUCES ANCHORAGE-INDEPENDENT CELL-CYCLE PROGRESSION

Normal fibroblasts are dependent on adhesion to a substrate for cell c ycle progression, Adhesion-deprived Rat1 cells arrest in the G(1) phas e of the cell cycle, with low cyclin E-dependent kinase activity, low levels of cyclin D1 protein, and high levels of the cyclin-dependent k inase inhibitor p27(kip1). To understand the signal transduction pathw ay undergoing adhesion-dependent growth, it is important to know wheth er prevention of any one of these down-regulation events under conditi ons of adhesion deprivation is sufficient to prevent the G(1) arrest, To that end, sublines of Rat1 fibroblasts capable of expressing cyclin E, cyclin D1, or both in an inducible manner were used, Ectopic expre ssion of cyclin D1 was sufficient to allow cells to enter S phase in a n adhesion-independent manner. In contrast, cells expressing exogenous cyclin E at a level high enough to overcome the p27(kip1)-imposed inh ibition of cyclin E-dependent kinase activity still arrested in G(1) w hen deprived of adhesion. Moreover, expression of both cyclins D1 and E in the same cells did not confer any additional growth advantage upo n adhesion deprivation compared to the expression of cyclin D1 alone. Exogenously expressed cyclin D1 was down-regulated under conditions of adhesion deprivation, despite the fact that it was expressed from a h eterozygous promoter. The ability of cyclin D1-induced cells to enter S phase in an adhesion independent manner disappears as soon as cyclin D1 proteins disappear. These results suggest that adhesion-dependent cell cycle progression is mediated through cyclin D1, at least in Rat1 fibroblasts.