Neutrophil extracellular trap formation requires OPA1-dependent glycolytic ATP production

Abstract

Optic atrophy 1 (OPA1) is a mitochondrial inner membrane protein that has an important role in mitochondrial fusion and structural integrity. Dysfunctional OPA1 mutations cause atrophy of the optic nerve leading to blindness. Here, we show that OPA1 has an important role in the innate immune system. Using conditional knockout mice lacking Opa1 in neutrophils (Opa1N∆), we report that lack of OPA1 reduces the activity of mitochondrial electron transport complex I in neutrophils. This then causes a decline in adenosine-triphosphate (ATP) production through glycolysis due to lowered NAD+ availability. Additionally, we show that OPA1-dependent ATP production in these cells is required for microtubule network assembly and for the formation of neutrophil extracellular traps. Finally, we show that Opa1N∆ mice exhibit a reduced antibacterial defense capability against Pseudomonas aeruginosa.

In vivo evidence for extracellular DNA trap formation

“…The conversion to a mucoid phenotype coincided with a decline in susceptibility to NETs, raising the possibility that increased alginate production decreases interactions with NETs, or otherwise interferes with killing by NET-associated granule proteins 48 . 4,6,7,144,152 . If the source of infection/stimulation persists, the released mtDNA, having similarity to viral and bacterial DNA (enriched in unmethylated CpG motifs), acts as a danger signal and triggers cytokine production for a protective and regulated immune response 7,55 .…”

“…4,6,7,144,152 . If the source of infection/stimulation persists, the released mtDNA, having similarity to viral and bacterial DNA (enriched in unmethylated CpG motifs), acts as a danger signal and triggers cytokine production for a protective and regulated immune response 7,55 . b Cytolysis: Under pathological conditions, such as the persistent presence of foreign antigens 104 , "too large to be trapped antigens", such as fungal hyphae 60,61 , strong adhesion receptor activation [153][154][155] , presence of monosodium urate (MSU) 88 , or phorbol-myristate-acetate (PMA) stimulation 2,12 , results in an excess of reactive oxygen species (ROS), leading to neutrophil cytolysis.…”

“…Moreover, optic atrophy 1 (OPA1), one of five GTPase dynamin family members, known to play a role in mitochondrial (mt) fusion, has recently been shown to be required for ATP production through glycolysis in neutrophils. If increases in ATP production are blocked, the assembly of the microtubule network and thus the formation of NETs do not occur 7 . ATP and ATP channel pannexin 1 (Panx1) contribute to NET formation and may represent therapeutic targets 8 .…”

Chemokine-triggered microtubule polymerization promotes neutrophil chemotaxis and invasion but not transendothelial migration

“…We found that different chemotactic peptides, including the key vascular chemokine CXCL1 and the end-target chemoattractant fMLP, triggered very slow but robust MT polymerization in these leukocytes, consistent with recent studies on fMLP-and C5a-triggered neutrophils. [25][26][27] Using a probe of real-time tubulin polymerization, SiR-tubulin, we found that neutrophils elongated their MTs within minutes when encountering these chemotactic peptides at their soluble states. Notably, rather than stabilizing these de novo-generated MTs, the MT binding drug taxol dramatically suppressed this slow chemoattractant-triggered MT polymerization.…”

“…In the present study, we have addressed these standing questions in a model of freshly isolated human neutrophils. We found that different chemotactic peptides, including the key vascular chemokine CXCL1 and the end‐target chemoattractant fMLP, triggered very slow but robust MT polymerization in these leukocytes, consistent with recent studies on fMLP‐ and C5a‐triggered neutrophils . Using a probe of real‐time tubulin polymerization, SiR‐tubulin, we found that neutrophils elongated their MTs within minutes when encountering these chemotactic peptides at their soluble states.…”