Old yellow enzyme protects the actin cytoskeleton from oxidative stress.
Old Yellow Enzyme (OYE) has long served as a paradigm for the study of flavin-containing NADPH oxido-reductases and yet its physiological role has remained a mystery. A two-hybrid interaction between Oye2p and actin led us to investigate a possible function in the actin cytoskeleton. We found that oye deletion strains have an overly elaborate actin cytoskeleton that cannot be attributed to changes in actin concentration but likely reflect stabilization of actin filaments, resulting in excessive actin assembly. Cells expressing the actin mutant act1-123p, which has a weakened interaction with Oye2p, show comparable defects in actin organization to the oye deletion strain that can be suppressed by overexpression of Oye2p. Similarly, mutation of either conserved cysteine of the potential disulfide pair Cys285-Cys374 in actin completely suppresses the actin organization defect of the oyeDelta phenotype. Strains lacking Oye function are also sensitive to oxidative stress as induced by H2O2, menadione, and diamide treatment. Mutation of either Cys285 or Cys374 of actin suppresses the sensitivity of oyeDelta strains to oxidative stress and in fact confers super-resistance to oxidative stress in otherwise wild-type strains. These results suggest that oxidative damage to actin, like that which has been observed in irreversibly sickled red blood cells, may be a general phenomenon and that OYE functions to control the redox state of actin thereby maintaining the proper plasticity of the actin cytoskeleton. In addition to uncovering a long sought biological function for Old Yellow Enzyme, these results establish that cellular sensitivity to oxidative stress can in part be directly attributed to a specific form (C285-C374 disulfide bond formation) of oxidative damage to actin.
Haarer
Brian K
BK
Department of Biochemistry and Molecular Biology, State University of New York, Upstate Medical University, Syracuse, NY 13210, USA.
Amberg
David C
DC
D000199:Actins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism*; D004220:Disulfides / Q000737:chemistry; D006801:Humans; D008958:Models, Molecular; D009154:Mutation; D009252:NADPH Dehydrogenase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D000199:Actins; D004220:Disulfides; D029701:Saccharomyces cerevisiae Proteins; D009252:NADPH Dehydrogenase