Human Ran cysteine 112 oxidation by pervanadate regulates its binding to keratins

J Biol Chem. 2005 Apr 1;280(13):12162-7. doi: 10.1074/jbc.M412505200. Epub 2005 Feb 3.

Abstract

We used a proteomic approach to identify proteins that associate with keratins 8 or 18 (K8/K18) in a pervanadate-dependent manner. Pervanadate triggers Ran-K8/K18 binding and a gel-migration-shift of Ran from 25 to 27 kDa, which does not occur upon exposure to H2O2 or vanadate or if pervanadate is excluded during cell solubilization. Generation of 27-kDa Ran is not related to hyperphosphorylation, is heat-insensitive, but occurs upon conversion of Ran cysteines to cysteic acid. The pervanadate-mediated Ran cysteine --> cysteic acid oxidation and its related gel migration shift affects other proteins including actin. Mutation of the three Ran cysteines (Cys-85, -112, and -120) showed that Ran Cys-112 oxidation generates 27-kDa Ran and accounts for its keratin binding. Proteasome inhibition accentuates Ran-keratin binding after cell exposure to pervanadate. Therefore, cell-free exposure to pervanadate causes cysteine to cysteic acid oxidation of Ran and several other proteins and Ran-K8/K18 association. In cells, stabilization of oxidized Ran by proteasome inhibition promotes Ran-keratin interaction. Keratin sequestration of oxidized Ran may provide a back-up protective mechanism in some cases of oxidative injury.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Actins / chemistry
  • Cell Line, Tumor
  • Cell Movement
  • Cell Survival
  • Cell-Free System
  • Cysteine / chemistry
  • DNA, Complementary / metabolism
  • Electrophoresis, Polyacrylamide Gel
  • Enzyme Inhibitors / pharmacology
  • Humans
  • Immunoprecipitation
  • Keratins / chemistry*
  • Mass Spectrometry
  • Models, Biological
  • Mutation
  • Oxygen / metabolism*
  • Phosphorylation
  • Proteasome Inhibitors
  • Protein Binding
  • Proteomics
  • Tyrosine / metabolism
  • Vanadates / pharmacology*
  • ran GTP-Binding Protein / chemistry*
  • ran GTP-Binding Protein / metabolism

Substances

  • Actins
  • DNA, Complementary
  • Enzyme Inhibitors
  • Proteasome Inhibitors
  • pervanadate
  • Vanadates
  • Tyrosine
  • Keratins
  • ran GTP-Binding Protein
  • Cysteine
  • Oxygen