EVIDENCE OF A FUNCTIONAL-ROLE FOR THE CYCLIN-DEPENDENT KINASE INHIBITOR P21(WAF1 CIP1/MDA6) IN THE RECIPROCAL REGULATION OF PKC ACTIVATOR-INDUCED APOPTOSIS AND DIFFERENTIATION IN HUMAN MYELOMONOCYTIC LEUKEMIA-CELLS/

functional role of the cyclin-dependent kinase inhibitor p21(WAF1/CIP1 ) in leukemic cell G(1) arrest, differentiation, and apoptosis induced by two PKC activators (PMA and bryostatin 1) was examined using antis ense-expressing lines [U937/p21AS(F4) and U937/p21AS(B8)]. Following i ncubation with 10 nM PMA (24 h), antisense-expressing cells displayed induction of p27(KIP1) but not of p21, whereas empty vector-containing cells (U937/pREP4) exhibited induction of both p21 and p27. Antisense -expressing cells were impaired in G(1) arrest, dephosphorylation of t he retinoblastoma protein, dephosphorylation and reduction in activity of cyclin-dependent kinase 2, and acquisition of differentiated featu res (e.g., plastic adherence). Bryostatin 1 induced p27 but not p21 in control cells and was less effective than PMA in initiating G(1) arre st and related events. Nevertheless, disruption of p21 expression abro gated the effects of bryostatin 1 on cell cycle arrest and cellular ma turation. Dysregulation of p21 did not, however, modify PMA- or bryost atin 1-mediated downregulation of c-Myc protein. Unexpectedly, disrupt ion of p21 failed to attenuate the net reduction in viable cell number following PMA or bryostatin 1 treatment inasmuch as impaired differen tiation was accompanied by a lowered threshold for PMA- and bryostatin 1-induced apoptosis. Inhibition of p21 expression also promoted PMA- and bryostatin 1-mediated loss of mitochondrial transmembrane potentia l (Delta Psi(m)) and release of cytochrome c into the cytosol. Togethe r, these findings demonstrate a critical functional role for p21 in re gulating myelomonocytic leukemic cell Gr, arrest and differentiation f ollowing exposure to two PKC activators exhibiting disparate patterns of activity. They also suggest that following treatment with these age nts, dysregulation of p21 prevents leukemic cells from engaging a norm al differentiation program through a c-Myc-independent mechanism, and instead directs cells along an apoptotic pathway. (C) 1998 Academic Pr ess.