Phorbol 12-myristate 13-acetate induces alteration in mucin gene expression and biological properties of colon cancer cells

Phorbol esters such as phorbol 12-myristate 13-acetate (PMA) have been repo rted to modulate diverse cellular responses through signal transduction pat hways including the protein kinase C (PKC) pathway. In the present study, w e sought to determine the effect of PMA on mucin gene expression and on the biological properties of a human colon cancer cell line, HM3. The cells we re treated for 8 and 24 h with various concentrations of PMA and total RNA was extracted and Northern and slot blot analyses were carried out using MU C2, MUC3 and MUC5AC mucin cDNA probes to assess the steady stare levels of mRNA. Spent media were collected and the level of cancer associated carbohy drate antigens (T, Tn, sialyl Tn, sialyl Le(x), and sialyl Le(a)) and matri x-degrading metalloproteinase (MMPs) activity were examined. Trypsinized ce lls were used for assessing in vitro invasion, motility and adhesion to mat rigel. Our results showed that PMA caused upregulation of steady state mRNA levels of MUC2, MUC3 and MUC5AC which was inhibited after treatment with p rotein synthesis inhibitors. Calphostin C, a highly specific inhibitor of p rotein kinase C significantly inhibited the PMA induced induction of mRNA l evels of MUC2, MUC3, and MUC5AC. The levels of all cancer-associated mucin carbohydrate antigens examined in the media were increased by PMA treatment . PMA also caused an increase in MMPs activity and in irt vitro invasion an d motility properties, but did not affect adhesion of HM3 cells to matrigel . Thus, PMA caused a significant increase in the expression of all three mu cin genes through signaling pathways involving protein kinase C and increas ed secretion of mucin associated carbohydrate antigens. These changes were associated with increases in MMP activity as well as by increases in the in vasive and motility properties of HM3 colon cancer cells. These data sugges t that protein kinase C signaling pathways may be involved in mucin gene re gulation and in modulating the invasive and metastatic properties of colon cancer cells.