Histone hypercitrullination mediates chromatin decondensation and neutrophil extracellular trap formation

TLDR

Neutrophils release highly decondensed chromatin structures called NETs, and while PAD4 is known to citrullinate histones, its role in neutrophil chromatin structure remains unclear. The study aimed to determine whether PAD4‑mediated histone hypercitrullination drives chromatin decondensation in neutrophils. Citrullination of defined nucleosome arrays by PAD4 blocks linker histone H5‑mediated compaction, preventing higher‑order chromatin formation. Hyper­citrullinated histones were found on decondensed chromatin in HL‑60 cells and neutrophils; PAD4 inhibition reduced citrullination and NET‑like structures, whereas PAD4 treatment promoted them, and loss of heterochromatin and multilobular nuclei was observed, indicating hypercitrullination is essential for chromatin decondensation in granulocytes.

Abstract

Peripheral blood neutrophils form highly decondensed chromatin structures, termed neutrophil extracellular traps (NETs), that have been implicated in innate immune response to bacterial infection. Neutrophils express high levels of peptidylarginine deiminase 4 (PAD4), which catalyzes histone citrullination. However, whether PAD4 or histone citrullination plays a role in chromatin structure in neutrophils is unclear. In this study, we show that the hypercitrullination of histones by PAD4 mediates chromatin decondensation. Histone hypercitrullination is detected on highly decondensed chromatin in HL-60 granulocytes and blood neutrophils. The inhibition of PAD4 decreases histone hypercitrullination and the formation of NET-like structures, whereas PAD4 treatment of HL-60 cells facilitates these processes. The loss of heterochromatin and multilobular nuclear structures is detected in HL-60 granulocytes after PAD4 activation. Importantly, citrullination of biochemically defined avian nucleosome arrays inhibits their compaction by the linker histone H5 to form higher order chromatin structures. Together, these results suggest that histone hypercitrullination has important functions in chromatin decondensation in granulocytes/neutrophils.