Oxidative stress induces S-thiolation of specific proteins in cultured gastric mucosal cells.
Oxidative stress induces the formation of protein-mixed disulfides with low-molecular-weight thiols, especially glutathione. We analyzed this process, termed S-thiolation, in cultured gastric mucosal cells from guinea pigs by gel electrophoresis and autoradiography after radiolabeling of the intracellular glutathione pool with 35S. Hydrogen peroxide (H2O2) or diamide initiated rapid and reversible S-thiolation of specific proteins with molecular masses of 42, 30, 29, 28, and 22 kDa. Diamide caused particularly prominent S-thiolation of the 42 kDa protein. This protein was identified as actin by immunoblot analysis and actin-myosin precipitation. Fluorescence microscopy revealed that diamide caused a disappearance of normal stress fibers and a concomitant increase in actin polymerization in association with contraction of the cells. These morphological changes were completely reversible within minutes. With cells depleted of glutathione by incubation with DL-buthionine-[S,R]-sulfoximine, diamide caused severe contraction and rounding, and the cells detached from the culture plates. S-thiolation of actin could help protect gastric mucosal cells against irreversible organization of microfilaments by preserving microfilament dynamics under oxidative stress.
Rokutan
K
K
Department of Nutrition, School of Medicine, University of Tokushima, Japan.
Johnston
R B
RB
Kawai
K
K
D008841:Actin Cytoskeleton / Q000648:ultrastructure; D000199:Actins / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D003553:Cystine / Q000493:pharmacokinetics; D003958:Diamide / Q000494:pharmacology*; D005753:Gastric Mucosa / Q000166:cytology / Q000378:metabolism* / Q000648:ultrastructure; D005978:Glutathione / Q000378:metabolism; D006168:Guinea Pigs; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D007425:Intracellular Membranes / Q000378:metabolism; D008297:Male; D008856:Microscopy, Fluorescence; D011506:Proteins / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000378:metabolism*
D000199:Actins; D011506:Proteins; D013438:Sulfhydryl Compounds; D003958:Diamide; D003553:Cystine; D006861:Hydrogen Peroxide; D005978:Glutathione