Nitration of PPARγ inhibits ligand-dependent translocation into the nucleus in a macrophage-like cell line, RAW 264

Abstract

Nitration of tyrosine residues in proteins has been observed in many inflammatory tissues of arthritis, ulcerative colitis, septic shock and ischemia-reperfusion injury. Although several studies have been carried out, it is still unclear what type of protein is nitrated and whether tyrosine nitration interferes with protein function. Peroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear receptor whose activation is linked to several physiological pathways including regulation of insulin sensitivity and control of inflammation. PPARγ possesses several tyrosine residues, which might be potential targets for nitration by peroxynitrite during inflammatory responses. Here we have investigated whether PPARγ is nitrated in macrophage-like RAW 264 cells and the effect of nitration on the translocation of PPARγ into the nucleus. Western blot analysis showed that tumor necrosis factor-α, lipopolysaccharide or peroxynitrite treatment significantly increases the nitration of PPARγ. Cell fractionation analysis and immunofluorescence coupled with confocal laser microscopy revealed that nitration of PPARγ inhibits its ligand-dependent translocation from the cytosol into the nucleus. Together, these results indicate that nitration of PPARγ during inflammation may be involved in a reduction in the control of inflammatory responses and also in the development of resistance to PPARγ ligand-based therapies against inflammation.