REGULATED EXPRESSION OF THE RETINOBLASTOMA SUSCEPTIBILITY GENE IN MAMMARY-CARCINOMA CELLS RESTORES CYCLIN D1 EXPRESSION AND G(1)-PHASE CONTROL

The product of the retinoblastoma susceptibility tumour suppressor gen e, pRb, is a negative regulator of cell proliferation. In order to inv estigate the interaction between pRb and the cell cycle machinery in m ore detail, a functional Rb gene was reintroduced into the Rb-deficien t human mammary carcinoma cell line BT549. Since constitutive high lev el expression of Rb turned out to be difficult to maintain, the tetrac ycline-dependent gene expression system was used. A number of clones w as generated which all showed low level expression in the noninduced s tate. Considerable induction rates were obtained. The low level of non induced Rb expression was sufficient to induce the expression of cycli n D1 the level of which was not further increased by upregulation of R b expression. Concomittantly, an increase in cell doubling time was ob served due to retardation of the cell cycle in the G1-phase. The data suggest that limiting amounts of cyclin D1 determine, at least partly, the extent of growth-repressing properties of pRb, The inducible syst em allows for maintainance of Rb-reconstituted cells at a low level of expression and for their use in the investigation of downstream funct ions of pRb.