Sulfhydration switch

Mol. Cell 45, 13–24 (2012)

Nuclear factor κB (NF-κB) is a transcription factor that acts downstream of tumor necrosis factor-α (TNF-α) and regulates antiapoptotic gene expression. Sen et al. now show that TNF-α signaling induces sulfhydration (R-SSH) of NF-κB and that this modification is critical for transcriptional activation. Hypothesizing that NF-κB activity may be linked to H₂S—a second messenger produced by cystathionine γ-lyase (CSE)—the authors showed that TNF-α treatment led to enhanced CSE and H₂S levels in cells and that DNA binding by NF-κB was dependent on CSE activity. Cysteine tagging and MS revealed that Cys38 within the DNA-binding p65 subunit of NF-κB underwent sulfhydration by H₂S. Cys38 had previously been identified as a site for S-nitrosylation that inhibits NF-κB activity, so the authors used a two-color tagging approach to show that Cys38 sulfhydration peaks shortly after TNF-α activation and that the amount of S -nitrosylcysteine is maximal at later times. The transcriptional activation profile of TNF-α–treated cells showed a similar time course, and additional biochemical analysis revealed that p65 sulfhydration enhances NF-κB binding to ribosomal protein S3 (RPS3), a known transcriptional coactivator. These results outline a time-dependent transcriptional regulatory mechanism by NF-κB that involves TNF-α induction of H₂S biosynthesis, selective p65 sulfhydration and transcriptional activation in collaboration with RPS3, which is followed by nitrosylation-dependent deactivation of gene expression.