Tyrosine phosphorylation and cellular redistribution of ezrin in MDCK cells treated with pervanadate.
Ezrin is a key protein in membrane-cytoskeleton interaction and is expressed primarily in actin-rich surface projections. Activation in protein tyrosine phosphorylation apparently regulates the structure and function of ezrin. In this study, we found that pervanadate (PV, the complexes of vanadate with hydrogen peroxide) caused an increase in tyrosine phosphorylation of ezrin and affected its cellular redistribution. Treatment of Madin-Darby canine kidney (MDCK) cells with pervanadate resulted in a dramatically increased tyrosine phosphorylation of ezrin within two to five min and the level reached the maximum after 60 min. This was accompanied by an alteration in the subcellular distribution of ezrin. Immunofluorescence and scanning laser confocal microscopy analysis revealed that, after PV stimulation, ezrin was redistributed from cytosol to the apical and lateral membrane domains. This occurred within five min, and more obvious redistribution to the lateral membrane domain was observed after 30 min. Furthermore, immunoblotting of ezrin in cell fractionation experiments showed that, in PV-treated MDCK cells, cytosolic ezrin was translocated to the membrane fraction, while there was no change in the level of ezrin associated with the actin-cytoskeleton. Therefore, cytoplasmic signaling may result in activation of ezrin in tyrosine phosphorylation, which is induced by PV stimulation. These results suggest that ezrin has qualities that might play a role in modulation of cell shape and adhesion.
Wu
Y X
YX
Department of Biochemistry, Hamamatsu University School of Medicine, 3600 Handa-cho, Hamamatsu-shi, Japan. yixinwu@hama-med.ac.jp
Uezato
T
T
Fujita
M
M
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000737:chemistry; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004285:Dogs; D007668:Kidney / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000276:immunology / Q000378:metabolism*; D010766:Phosphorylation; D014443:Tyrosine / Q000378:metabolism*; D014638:Vanadates / Q000494:pharmacology*; D000099149:Ezrin
D000906:Antibodies; D003598:Cytoskeletal Proteins; D010750:Phosphoproteins; D014443:Tyrosine; D014638:Vanadates; D000099149:Ezrin; C076715:pervanadate