Independent regulation of adherens and tight junctions by tyrosine phosphorylation in Caco-2 cells

To study the role of tyrosine phosphorylation in the control of intercellul ar adhesion of intestinal cells, we have generated several clones of Caco-2 cells that express high levels of pp60v-src only after addition of butyrat e. Expression of this oncogene in cells 5 days after confluence induced bet a-catenin and p120-ctn tyrosine phosphorylation, redistribution of E-cadher in to the cytosol and disassembly of adherens junctions. However, tight jun ctions of Caco-2 cells at 5 days after confluence were not altered by expre ssion of pp60v-src. Similar results were obtained when Caco-2 cells were in cubated with phosphotyrosine phosphatase inhibitor orthovanadate. Although addition of this compound to postconfluent cells disrupt adherens junctions , tight junctions remain unaltered, as determined measuring monolayer perme ability to mannitol or hyperphosphorylation of Triton-insoluble occludin. M odifications in tight junction permeability of Caco-2 were only observed at high concentrations of orthovanadate (1 mM). Interestingly, this tyrosine phosphorylation-refractory state was achieved after confluence since early postconfluent cells (day 2) showed a limited but significant response to lo w doses of orthovanadate. These results suggest that tight junctions of dif ferentiated Caco-2 cells are uncoupled from adherens junctions and are inse nsitive to regulation by tyrosine phosphorylation. (C) 1999 Elsevier Scienc e B.V. All rights reserved.