Ectopic expression of Cdk6 circumvents transforming growth factor-beta mediated growth inhibition

Transforming growth factor-beta (TGF-beta) induced growth arrest of cells i nvolves regulation of the activities of both D- and E-type cyclin kinase co mplexes thought to be mediated primarily by the regulation of p15(Ink4b) an d p27(Kip1) cyclin kinase inhibitors. We show here that TGF-beta downregula tes Cdk6 and that transient and stable expression of Cdk6 in Mv1Lu mink epi thelial cells overrides TGF-beta mediated arrest. The main effect of the ec topic Cdk6 expression was to sequester TGF-P induced p15(Ink4b) and to main tain more p27(Kip1) in cyclin D-complexes preventing the complete shift of p27(Kip1) to Cdk2 invoked by TGF-beta. This led to the presence of an activ e cyclinD-Cdk6-p27(Kip1) complex and partially active cyclin E-Cdk2 complex and resulted in the failure of TGF-beta to fully arrest Mv1Lu cell growth. Though dominant negative Cdk6, expressed similarly in the cells, sequester ed both p15(Ink4b) and p27(Kip1), it lacks kinase activity and was unable t o override the TGF-beta arrest. The results demonstrate that downregulation of Cdk6 kinase is required for the enforcement of the G(1)-phase arrest by TGF-beta and results in changes in association of the p15(Ink4b) and p27(K ip1) inhibitors with D- and E-type cyclin kinase complexes.