POST-PROLIFERATIVE CYCLIN E-ASSOCIATED KINASE-ACTIVITY IN DIFFERENTIATED OSTEOBLASTS - INHIBITION BY PROLIFERATING OSTEOBLASTS AND OSTEOSARCOMA CELLS

Spontaneous differentiation of normal diploid osteoblasts in culture i s accompanied by increased cyclin E associated kinase activity on (1) the retinoblastoma susceptibility protein pRB, (2) the p107 RE related protein, and (3) two endogenous cyclin E-associated substrates of 78 and 105 kD. Activity of the differentiation-related cyclin E complexes (diff.ECx) is not recovered in cdc2 or cdk2 immunoprecipitates. Phosp horylation of both the 105 kD endogenous substrate and the p107 exogen ous substrate is sensitive to inhibitory activity (diff.ECx-i) present in proliferating osteoblasts. This inhibitory activity is readily rec ruited by the cyclin E complexes of differentiated osteoblasts but is not found in cyclin E immunoprecipitates of the proliferating eel Is t hemselves. Strong inhibitory activity on diff.ECx kinase activity is e xcerted by proliferating ROS 17/2.8 osteosarcoma cells. However, unlik e the normal diploid cells, the diff. ECx-i activity of proliferating ROS 1 7/2.8 cells is recovered by cyclin E immunoprecipitation. The cy clin-dependent kinase inhibitor p21(CIP1/WAF1) inhibits diff. ECx kina se activity. Thus, our results suggest the existence of a unique regul atory system, possibly involving p21(CIP1/WAF1), i, which inhibitory a ctivity residing in proliferating cells is preferentially targeted tow ards differentiation-related cyclin E-associated kinase activity. (C) 1997 Wiley-Liss, Inc.