PROTEIN-KINASE-C ISOZYME-MEDIATED CELL-CYCLE ARREST INVOLVES INDUCTION OF P21(WAF1 CIP1) AND P27(KIP1) AND HYPOPHOSPHORYLATION OF THE RETINOBLASTOMA PROTEIN IN INTESTINAL EPITHELIAL-CELLS/

The molecular mechanisms underlying protein kinase C (PKC) isozyme-med iated control of cell growth and cell cycle progression are poorly und erstood. Our previous analysis of PKC isozyme regulation in the intest inal epithelium in situ revealed that multiple members of the PKC fami ly undergo changes in expression and subcellular distribution precisel y as the cells cease proliferating in the mid-crypt region, suggesting that activation of one or more of these molecules is involved in nega tive regulation of cell growth in this system (Saxon, M. L., Zhao, X., and Black, J. D. (1994) J. Cell Biol. 126, 747-763). In the present s tudy, the role of PKC isozyme(s) in control of intestinal epithelial c ell growth and cell cycle progression was examined directly using the IEC-18 immature crypt cell line as a model system. Treatment of IEC-18 cells with PKC agonists resulted in translocation of PKC alpha, delta , and epsilon from the soluble to the particulate subcellular fraction , cell cycle arrest in G(1) phase, and delayed transit through S and/o r G(2)/M phases. PKC-mediated cell cycle arrest in G(1) was accompanie d by accumulation of the hypophosphorylated, growth-suppressive form o f the retinoblastoma protein and induction of the cyclin-dependent kin ase inhibitors p21(waf1/cip1) and p27(kip1). Reversal of these cell cy cle regulatory effects was coincident with activator-induced down-regu lation of PKC alpha, delta, and epsilon. Differential down-regulation of individual PKC isozymes revealed that PKC alpha in particular is su fficient to mediate cell cyclearrest by PKC agonists in this system. T aken together, the data implicate PKC alpha in negative regulation of intestinal epithelial cell growth both in vitro and in situ via pathwa ys which involve modulation of Cip/Kip family cyclin-dependent kinase inhibitors and the retinoblastoma growth suppressor protein.