EVIDENCE THAT TYROSINE PHOSPHORYLATION MAY INCREASE TIGHT JUNCTION PERMEABILITY

Tight junction permeability control is important in a variety of physi ological and pathological processes, We have investigated the role of tyrosine phosphorylation in the regulation of tight junction permeabil ity. MDCK epithelial cells and brain endothelial cells were grown on f ilters and tight junction permeability was determined by transcellular electrical resistance (TER), The tyrosine phosphatase inhibitor perva nadate caused a concentration- and time-dependent decrease in TER in b oth MDCK and brain endothelial cells, However, as expected, pervanadat e resulted in the tyrosine phosphorylation of many proteins; hence int erpretation of its effects are extremely difficult, Phenylarsine oxide , a more selective tyrosine phosphatase inhibitor, caused the tyrosine phosphorylation of relatively few proteins as analyzed by immunoblott ing of whole cell lysates, This inhibitor, like pervanadate, also elic ited a decrease in TER in the two cell types, In the MDCK cells, the a ction of phenylarsine oxide could be reversed by the subsequent additi on of the reducing agent 2,3-dimercaptopropanol. Immunocytochemistry r evealed that phenylarsine oxide rapidly stimulated the tyrosine phosph orylation of proteins associated with intercellular junctions. Because of the known influence of the adherens junction on tight junctions, w e analyzed immunoprecipitates of the E-cadherin/catenin complex from M DCK cells treated with phenylarsine oxide. This revealed an increase i n the tyrosine phosphorylation of beta-catenin, but not of alpha-caten in. However, the tight junction associated protein ZO-1 was also tyros ine phosphorylated after PAO treatment, These data indicate that tight junction permeability may be regulated via mechanisms involving tyros ine phosphorylation of adherens junction and tight junction proteins.