Targeted degradation of extracellular mitochondrial aspartyl-tRNA synthetase modulates immune responses in bacterial pneumonia

The severity of bacterial pneumonia can be worsened by impaired innate immunity resulting in ineffective pathogen clearance. We describe a mitochondrial protein, aspartyl-tRNA synthetase (DARS2), which is released in circulation during bacterial pneumonia in humans and displays intrinsic innate immune properties and cellular repair properties. DARS2 interacts with a bacterial-induced ubiquitin E3 ligase subunit, FBXO24, which targets DARS2 for ubiquitylation and degradation, a process that is inhibited by DARS2 acetylation. During experimental pneumonia, Fbxo24 knockout mice exhibit elevated DARS2 levels with a robust increase in pulmonary cellular and cytokine levels. In silico modeling identified an FBXO24 inhibitory compound with immunostimulatory properties which extended DARS2 lifespan in cells. Here we show a unique biological role for an extracellular, mitochondrially derived enzyme and its molecular control by the ubiquitin apparatus, which may serve as a mechanistic platform to enhance protective host immunity through small molecule discovery. Overall design: BEAS-2B Cells stably integrated with CRSPR/Cas9 and gRNA scramble (sgCon) or against FBXO24 (sgO24). sgCon and sgO24 cells were infected with PA103 MOI1 or left uninfected for 24h. Cells were collected andprocessed for RNAseq to determine differences in cellular pathways.