Histones in neutrophil extracellular traps (NETs) contain oxidative post-translational modifications induced by the myeloperoxidase oxidant hypochlorous acid (HOCl)

ABSTRACT: Extracellular traps (NETs) are released by neutrophils during inflammation and play a role in pathology of diseases such as atherosclerosis, as well as their innate immune function of clearing infection. NETs are composed of a mesh of DNA and histones and contain various anti-microbial granule proteins, such as myeloperoxidase (MPO), which remains enzymatically active. This can result in the production of the reactive oxidant hypochlorous acid (HOCl), which reacts readily with proteins. However, whether histones and other NET-associated proteins contain HOCl-modified proteins is unknown. This is significant as recent studies show that exposure of histones to HOCl alters their extracellular reactivity to promote inflammatory signaling in vascular cells. In this study, we used a proteomic approach to characterise the protein composition of NETs and examine whether histones and other proteins contain oxidative protein-translational modifications resulting from reactions with HOCl. NETs were collected from neutrophil-like PLB-985 cells and primary neutrophils stimulated with phorbol myristate acetate (PMA) or nigericin, a bacterial peptide derived from Streptomyces hygroscopicus. Exposure of proteins to HOCl results in the formation of nitrile and aminoadipic semialdehyde formation on lysine residues, chlorination of tyrosine and tryptophan residues together with methionine oxidation products. Evidence was obtained for the presence of tyrosine chlorination on histones, particularly histone H4, and other proteins like quinone oxidoreductase. Other PTMs, including citrullination and acetylation were also detected on histones (H1, H2A, H3 and H4). Chlorination of the histone H4 peptide TVTAMDVVYALK(R) by HOCl was more abundant in NETs obtained from neutrophils exposed to PMA compared to nigericin, consistent with nigericin triggering NET release via a non-oxidative pathway. Current studies are investigating whether histone modifications occur before or after release from the cell. These findings provide the first evidence that HOCl induces histone modifications on NETs, which may be relevant in disease.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Neutrophil

DISEASE(S): Inflammation

SUBMITTER: Per Hägglund  

LAB HEAD: Clare Hawkins and Per Hägglund

PROVIDER: PXD060348 | Pride | 2025-05-30

REPOSITORIES: Pride