Unopposed phosphatase action initiates ezrin dysfunction: a potential mechanism for anoxic injury.
Because extensive kinase inhibition during anoxia has previously been reported, we investigated the role of kinase inhibition in anoxic cell injury by studying the effects of kinase inhibitors on a membrane-microvillar cytoskeleton linker protein, ezrin, in rabbit renal proximal tubules. Like anoxia, kinase inhibitors caused ezrin dephosphorylation in a dose-dependent manner under normoxia. The kinase inhibitor chelerythrine also induced ezrin dissociation from the cytoskeleton, i.e., causing it to lose its membrane-cytoskeleton linker function. Blockage of kinase inhibitor-induced ezrin dephosphorylation by a phosphatase inhibitor, calyculin A, ameliorated ezrin dissociation. Stimulation of the kinase during anoxia did not improve ezrin phosphorylation, suggesting that anoxia-induced kinase inhibition might be due to the lack of the substrate ATP. Finally, in vitro study of ezrin phosphatase revealed no increase in its activity during anoxia, suggesting the principal role of kinase inhibition in the loss of the linker function of ezrin during anoxia. Our results provide, for the first time at the molecular level, a mechanistic insight into anoxic cell injury caused by unopposed phosphatase action.
Chen
J
J
Department of Cell Biology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Mandel
L J
LJ
D000470:Alkaloids; D000818:Animals; D053119:Benzophenanthridines; D003598:Cytoskeletal Proteins; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D000860:Hypoxia / Q000473:pathology* / Q000503:physiopathology*; D007687:Kidney Tubules, Proximal / Q000201:enzymology; D008387:Marine Toxins; D010080:Oxazoles / Q000494:pharmacology; D010617:Phenanthridines / Q000494:pharmacology; D010750:Phosphoproteins / Q000502:physiology*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D010766:Phosphorylation; D010770:Phosphotransferases / Q000037:antagonists & inhibitors; D011493:Protein Kinase C / Q000378:metabolism; D011817:Rabbits; D012016:Reference Values; D000099149:Ezrin
D000470:Alkaloids; D053119:Benzophenanthridines; D003598:Cytoskeletal Proteins; D004791:Enzyme Inhibitors; D008387:Marine Toxins; D010080:Oxazoles; D010617:Phenanthridines; D010750:Phosphoproteins; D010744:Phosphoric Monoester Hydrolases; D010770:Phosphotransferases; D011493:Protein Kinase C; D000099149:Ezrin; C059041:calyculin A; C016299:chelerythrine