2. Laboratory of Signal Transduction, Hoshi University. 2-4-41 Ebara, Shinagawa-ku, Tokyo 142-8501, Japan.
We sought to determine how hyperactivation of p38 MAP kinase affects cell-cell adhesion in two epithelial cell lines. The first cell line, HCC2998/MKK6DA, a human colon adenocarcinoma cell line containing inducible dominant-active MKK6, has been shown to disperse after hyperactivation of p38 MAP kinase. However, the mechanism of how these cells disperse has not been studied. Examination of adherens junction proteins, such as E-cadherin and β -catenin, revealed that adherens junctions were disrupted. HCC2998/MKK6DA cells respond to heregulin β -1. MCF7, human breast cancer cells which also respond to heregulin β -1, were also examined for localization. MCF7/MKK6DA-6-2-4 cells carrying the inducible dominant-active MKK6 gene were produced. These cells also dispersed after hyperactivation of p38 MAP kinase. As these cells grow as monolayers, observation of the cells was more convenient than in HCC2998 cells. Dissociation of MCF7/MKK6DA-6-2-4 cells occurred 21–25 h after infection with adenovirus coding for Cre recombinase.. F-actin, which associates with the adherens junction, disappeared before cells dispersed. Then, E-cadherin and occludin disappeared from cell-cell junctions to localize diffusely in the cytosol, which led to loss of cell-cell interaction. This was different from the case with HCC2998 cells, in which E-cadherin and β -catenin remained at the plasma membrane after dissociation. Treatment with SB202190, a p38 MAP kinase inhibitor, blocked dissociation of the cells, indicating that p38 MAP kinase activity was required for cell dispersal. Localization of adherens junction proteins was examined in MCF7/MKK6-6-2-4 cells. Cells were fractionated into the cytosol and membrane fractions. After dissociation of the cells, E-cadherin remained in the membrane fraction; however, α -catenin and β -catenin moved from the membrane fraction to the cytosolic fraction, suggesting that these proteins dissociate from E-cadherin. Finally, activation of Rac1 , which has been shown to be important in dissociation of the cells was detected. These results suggest that hyperactivation of p38 MAP kinase causes dissociation of the cells through disruption of the adherens junction in these cell lines.
Keywords: p38 MAP kinase, adherens junction, cell-cell interaction, scattering