ZDHHC5-mediated S-palmitoylation of NOD2 drives neutrophil extracellular trap formation via mitophagy to exacerbate inflammation in COPD.

Abstract

Chronic obstructive pulmonary disease (COPD) is characterized by persistent neutrophilic inflammation, but its upstream molecular regulators remain incompletely defined. Here, NOD2 and the palmitoyl-acyltransferase ZDHHC5 were increased in COPD lung tissue and detected in pulmonary neutrophils. In CS-exposed mice, NOD2 deficiency attenuated lung inflammation, collagen deposition, EMT-associated changes, and neutrophil extracellular trap (NET) formation. Anti-Ly6G-mediated neutrophil depletion reproduced much of this protection and markedly reduced the differences between wild-type and NOD2-deficient mice, indicating that NOD2-associated pathology depends largely on neutrophils. In CSE-stimulated HL-60-derived neutrophil-like cells and primary mouse neutrophils, NOD2 loss reduced NET formation, intracellular reactive oxygen species accumulation, and mitophagy, whereas the autophagy inhibitor 3-MA attenuated NOD2-overexpression-induced NET formation. CSE also increased NOD2 S-palmitoylation. ZDHHC5 was associated with NOD2 and positively regulated its protein abundance, whereas mutation of C395 and C1033 reduced NOD2 palmitoylation, stability, membrane localization, and downstream signaling. ZDHHC5 overexpression partially counteracted the effects of NOD2 loss in vitro and in vivo, although residual effects in NOD2^-/- neutrophils indicated additional NOD2-independent mechanisms. Collectively, these findings identify ZDHHC5-NOD2 signaling as an important regulator of neutrophil-associated inflammation and NET formation in COPD.