LOSS OF RB AND MYC ACTIVATION COOPERATE TO SUPPRESS CYCLIN D1 AND CONTRIBUTE TO TRANSFORMATION

Cyclin D1 can bind and phosphorylate the product (pRb) of the retinobl astoma gene (RB-1) and recent evidence suggests pRb, in turn, may regu late cyclin D1 protein expression. In transformed cell lines, loss of pRb activity strongly correlates with a decrease in cyclin D1 protein expression, and conversely, introduction of pRb can induce cyclin D1 p romoter activity. We show here that pRb does not regulate cyclin D1 di rectly as basal and serum-stimulated levels of cyclin D1 protein and k inase activity are similar in wildtype and pRb-deficient primary mouse embryonic fibroblasts (MEFs). These observations suggest that the sup pression of cyclin D1 in pRb-minus tumour cell lines requires both los s of pRb and at least one additional genetic event. We have determined that constitutive, ectopic Myc expression in pRb-deficient, but not w ildtype, MEFs suppresses cyclin D1 protein expression and kinase activ ity. Regulation is evident at either the level of RNA or protein expre ssion. Phenotypically, pRb-deficient MEFs consistently exhibited a del ayed growth response in comparison to wildtype MEFs. This growth delay is abrogated in pRb-deficient MEFs which are expressing ectopic Myc p rotein, coincident with the loss of cyclin D1 protein expression. More over, these cells exhibit an increased proliferative capacity, and the y no longer show contact inhibition. Our results support a cross-regul atory mechanism between Myc, pRb and cyclin D1 and suggest a novel rol e for cyclin D1 in tumorigenesis.