The phosphatase inhibitor okadaic acid induces AQP2 translocation independently from AQP2 phosphorylation in renal collecting duct cells

Phosphorylation by kinases and dephosphorylation by phosphatase markedly af fect the biological activity of proteins involved in intracellular signalin g. In this study we investigated the effect of the serine/threonine phospha tase inhibitor okadaic acid on water permeability properties and on aquapor in2 (AQP2) translocation in AQP2-transfected renal CD8 cells. In CD8 cells both forskolin alone and okadaic acid alone increased the osmotic water per meability coefficient P-f by about 4- to 5-fold. In intact cells, in vivo p hosphorylation studies revealed that forskolin stimulation resulted in a th reefold increase in AQP2 phosphorylation. In contrast, okadaic acid treatme nt promoted only a 60% increase in AQP2 phosphorylation which was abolished when this treatment was performed in the presence of 1 mu M H89, a specifi c protein kinase A (PKA) inhibitor, Nevertheless, in this latter condition, confocal microscopy analysis revealed that AQP2 translocated and fused to the apical membrane. Okadaic acid-induced AQP2 translocation was dose depen dent having its maximal effect at a concentration of 1 mu M. In conclusion, our results clearly indicate that okadaic acid exerts a full forskolin-lik e effect independent from AQP2 phosphorylation. Thus AQP2 phosphorylation i s not essential for water channel translocation in renal cells, indicating that different pathways might exist leading to AQP2 apical insertion and in crease in P-f.