THE ENTEROCYTE-LIKE DIFFERENTIATION OF THE CACO-2 TUMOR-CELL LINE STRONGLY CORRELATES WITH RESPONSIVENESS TO CAMP AND ACTIVATION OF KINASE-A PATHWAY

We have investigated the expression of protein kinase C (PKC) and prot ein kinase A (PKA) during the phases of growth and differentiation of the human colon carcinoma Caco-2 cells. We studied whether differentia tion correlated with the responsiveness to cAMP and with an increased transport of the catalytic subunit of PKA into the nucleus. Also, we e valuated whether this phenomenon was affected by PKC activity. High le vels of activated PKC were found in the plasma membranes of replicatin g cells. When the cells began to differentiate, plasma membrane-activa ted PKC decreased, while the cytosolic fraction increased. On the cont rary, PKA holoenzyme increased during differentiation, along with the transport of its catalytic subunit into the nucleus. Both types I and II kinase A holoenzymes increased during differentiation, with maximal type II activity found when cells were fully differentiated. In repli cating preconfluent cells, the inhibition of PKC by high dose phorbol 12-myristate 13-acetate or sphingosine increased the amount of both PK A catalytic subunit in the nucleus and sucrase activity. During differ entiation, 8-Bromo-cAMP increased PKA catalytic subunit in the nucleus and apoliprotein A1 mRNA levels. These effects were inhibited by low- dose phorbol 12-myristate 13-acetate, which activates PKC in the plasm a membranes. Our data suggest that PKC is activated in proliferating C aco-2 cells. The inhibition of PKC induces the transport of PKA cataly tic subunit into the nucleus and the expression of the differentiation markers. Differentiated Caco-2 cells show a lower activation of PKC a nd an increased transport of the catalytic subunit of PKA into the nuc leus. Differentiated Caco-2 cells are highly responsive to cAMP and th e 8-Br-cAMP analog is able to revert the inhibitory effect of PKC.