Peptidyl arginine deiminase-4 (PAD4) catalyzes the conversion of peptidylarginine residues to peptidylcitrulline. We have previously shown that kidney ischemia-reperfusion (I/R) injury increases renal proximal tubular PAD4 expression and activity. Furthermore, kidney PAD4 plays a critical role in ischemic acute kidney injury (AKI) by promoting renal tubular inflammation, neutrophil infiltration, and NF-κB activation. However, the mechanisms of PAD4-mediated renal tubular inflammation and NF-κB activation after I/R remain unclear. Here, we show that recombinant PAD4 preferentially citrullinates recombinant IKKγ [also called NF-κB essential modulator (NEMO)] over recombinant IKKα or IKKβ. Consistent with this finding, PAD4 citrullinated renal proximal tubular cell IKKγ and promoted NF-κB activation via IκBα phosphorylation in vitro. NEMO inhibition with a selective NEMO-binding peptide attenuated PAD4-mediated proinflammatory cytokine mRNA induction in HK-2 cells. Moreover, NEMO inhibition did not affect proximal tubular cell survival, proliferation, or apoptosis, unlike global NF-κB inhibition. In vivo, NEMO-binding peptide treatment protected against ischemic AKI. Finally, NEMO-binding peptide attenuated recombinant PAD4-mediated exacerbation of ischemic AKI, renal tubular inflammation, and apoptosis. Taken together, our results show that PAD4 exacerbates ischemic AKI and inflammation by promoting renal tubular NF-κB activity and inflammation via NEMO citrullination. Targeting NEMO activation may serve as a potential therapy for this devastating clinical problem.
Keywords: NF-κB essential modulator; acute kidney injury; apoptosis; citrullination; inflammation; ischemia; necrosis; neutrophil; reperfusion.