Decrease in protein tyrosine phosphorylation is associated with F-actin reorganization by retinoic acid in human endometrial adenocarcinoma (RL95-2) cells

Transformed cells often express elevated levels of tyrosine-phosphorylated proteins. Inhibition of protein tyrosine kinases causes reversion of malign ant cells to the normal phenotype. In the present study, we evaluated the p ossibility that the reversion of human endometrial adenocarcinoma RL95-2 ce lls to a stationary phenotype induced by retinoic acid was associated with inhibition of tyrosine phosphorylation of cellular proteins. We found that retinoic acid decreased the levels of tyrosine-phosphorylated proteins, as assessed by immunostaining and immunoprecipitations using specific anti-pho sphotyrosine antibodies. In addition, the inhibitors of tyrosine kinases he rbimycin A and tyrphostin mimicked retinoic acid, inducing F-actin reorgani zation and increasing the size of RL95-2 cells, as determined by measuremen t of cell perimeters. Because focal adhesions that connect actin filaments with the plasma membrane are major sites of tyrosine phosphorylation, we fu rther investigated whether selected focal adhesion proteins were affected b y retinoic acid. We found that retinoic acid altered the localization of fo cal adhesion kinase. All-trans retinoic acid was effective in reducing the levels of focal adhesion kinase and paxillin protein. Thirteen-cis retinoic acid increased the levels of vinculin protein in the cytosolic fraction of cells. These changes are consistent with actin reorganization and reversio n toward a stationary phenotype induced by retinoic acid in endometrial ade nocarcinoma RL95-2 cells. Our results indicate that the differentiating eff ects of retinoids on endometrial cells are associated with decreases in tyr osine phosphorylation and changes in the levels and distribution of focal a dhesion proteins. These findings suggest that signaling pathways that invol ve tyrosine kinases are potential targets for drug design against endometri al cancer. J. Cell. Physiol. 178:320-332, 1999. (C) 1999 Wiley-Liss, Inc.