S-nitrosylation attenuates pregnane X receptor hyperactivity and acetaminophen-induced liver injury

Drug-induced liver injury (DILI), especially acetaminophen overdose, is the leading cause of acute liver failure. Pregnane X receptor (PXR) is a nuclear receptor and the master regulator of drug metabolism. Aberrant activation of PXR plays a pathogenic role in the acetaminophen hepatotoxicity. Here, we aimed to examine the S-nitrosylation of PXR (SNO-PXR) in response to acetaminophen. We found that PXR was S-nitrosylated in hepatocytes and the mouse livers after exposure to acetaminophen or S-nitrosoglutathione (GSNO). Mass spectrometry and sitedirected mutagenesis identified the cysteine 307 as the primary residue for S-nitrosylation (SNO) modification. In hepatocytes, SNO suppressed both agonist-induced (rifampicin and SR12813) and constitutively active PXR (VP-PXR, a human PXR fused to the minimal transactivator domain of the herpes virus transcription factor VP16) activations. Furthermore, in acetaminophenoverdosed mouse livers, PXR protein was decreased at the centrilobular regions overlapping with increased SNO. In PXR^-/- mice, replenishing the livers with the SNO-deficient PXR significantly aggravated hepatic necrosis, increased HMGB1 release, and exacerbated liver injury and inflammation. Particularly, we demonstrated that S-nitrosoglutathione reductase (GSNOR) inhibitor N6022 promoted hepatoprotection by increasing the levels of SNO-PXR. In conclusion, PXR is posttranslationally modified by SNO in hepatocytes in response to acetaminophen. This modification mitigated the acetaminophen-induced PXR hyperactivity. It may serve as a target for therapeutical intervention.