PLC-gamma 1 signaling pathway and villin activation are involved in actin cytoskeleton reorganization induced by Na+/P-i cotransport up-regulation

Background: The brief incubation of opossum kidney (OK) cells with low P-i results in Na+/P-i cotransport up-regulation and in substantial, but transi ent, cytoskeletal reorganization. In this study, we examined signaling even ts involved in the depolymerization of microfilaments. Results: Confocal laser scanning microscopy, immunoblot and immunoprecipita tion experiments revealed villin co-localization with mainly actin short fi laments and monomers, indicating that under the conditions used, villin act ed as an actin-severing protein. Further analysis revealed that low concent rations of extracellular phosphate resulted in phospholipase C gamma 1 (PLC -gamma 1) translocation to the actin cytoskeleton, without increases in its tyrosine phosphorylation. Additionally, tyrosine phosphorylation of a port ion of insoluble villin was increased; whereas, only tyrosine phosphorylate d villin associated with PLC-gamma 1. Although, tyrosine phosphorylation of PLC-gamma 1 was not observed during Na+/P-i cotransport up-regulation, gen istein treatment abolished the enzyme's translocation to the actin cytoskel eton, as well as its association with villin. In addition, villin was found to associate with the 85-KDa subunit (p85) of phosphatidylinositol (PI)-3 kinase, concomitant with PLC-gamma 1, in the cytoskeletal fraction of Na+/P -i cotransport up-regulated cells. Conclusions: Our observations suggest a signaling mechanism linking low amb ient P-i levels to the acute up-regulation of its cotransport with sodium a nd the depolymerization of the subcortical actin cytoskeleton.