ACTIVATION OF PROTEIN-KINASE-C ISOZYMES IS ASSOCIATED WITH POSTMITOTIC EVENTS IN INTESTINAL EPITHELIAL-CELLS IN-SITU

The mechanisms underlying control of cell growth and differentiation i n epithelial tissues are poorly understood. Protein kinase C (PKC) iso zymes, members of a large family of serine/threonine kinases of fundam ental importance in signal transduction, have been increasingly implic ated in the regulation of cell growth, differentiation, and function. Using the rat intestinal epithelium as a model system, we have examine d PKC-specific activity as well as individual PKC isozyme expression a nd distribution (i.e., activation status) in epithelial cells in situ. Increased PKC activity was detected in differentiating and functional cells relative to immature proliferating crypt cells. Immunofluoresce nce and Western blot analysis using a panel of isozyme-specific antibo dies revealed that PKC alpha, beta II, delta, epsilon, and zeta are ex pressed in rat intestinal epithelial cells and exhibit distinct subcel lular distribution patterns along the crypt-villus unit. The combined morphological and biochemical approach used permitted analysis of the activation status of specific PKC isozymes at the individual cell leve l. These studies showed that marked changes in membrane association an d level of expression for PKC alpha, beta II, delta, and zeta occur as cells cease division in the mid-crypt region and begin differentiatio n. Additional changes in PKC activation status are observed with acqui sition of mature function on the villus. These studies clearly demonst rate naturally occurring alterations in PKC isozyme activation status at the individual cell level within the context of a developing tissue . Direct activation of PKC in an immature intestinal crypt cell line w as shown to result in growth inhibition and coincident translocation o f PKC alpha from the cytosolic to the particulate subcellular fraction , paralleling observations made in situ and providing further support for a role of intestinal PKC isozymes in post-mitotic events. PKC isoz ymes were also found to be tightly associated with cytoskeletal elemen ts, suggesting participation in control of the structural organization of the enterocyte. Taken together, the results presented strongly sug gest an involvement of PKC isoforms in cellular processes related to g rowth cessation, differentiation, and function of intestinal epithelia l cells in situ.