RLR-mediated antiviral innate immunity requires oxidative phosphorylation activity

Scientific Reports, Jul 2017

Mitochondria act as a platform for antiviral innate immunity, and the immune system depends on activation of the retinoic acid-inducible gene I (RIG-I)-like receptors (RLR) signaling pathway via an adaptor molecule, mitochondrial antiviral signaling. We report that RLR-mediated antiviral innate immunity requires oxidative phosphorylation (OXPHOS) activity, a prominent physiologic function of mitochondria. Cells lacking mitochondrial DNA or mutant cells with respiratory defects exhibited severely impaired virus-induced induction of interferons and proinflammatory cytokines. Recovery of the OXPHOS activity in these mutants, however, re-established RLR-mediated signal transduction. Using in vivo approaches, we found that mice with OXPHOS defects were highly susceptible to viral infection and exhibited significant lung inflammation. Studies to elucidate the molecular mechanism of OXPHOS-coupled immune activity revealed that optic atrophy 1, a mediator of mitochondrial fusion, contributes to regulate the antiviral immune response. Our findings provide evidence for functional coordination between RLR-mediated antiviral innate immunity and the mitochondrial energy-generating system in mammals.

A PDF file should load here. If you do not see its contents the file may be temporarily unavailable at the journal website or you do not have a PDF plug-in installed and enabled in your browser.

Alternatively, you can download the file locally and open with any standalone PDF reader:

https://www.nature.com/articles/s41598-017-05808-w.pdf

RLR-mediated antiviral innate immunity requires oxidative phosphorylation activity

Abstract Mitochondria act as a platform for antiviral innate immunity, and the immune system depends on activation of the retinoic acid-inducible gene I (RIG-I)-like receptors (RLR) signaling pathway via an adaptor molecule, mitochondrial antiviral signaling. We report that RLR-mediated antiviral innate immunity requires oxidative phosphorylation (OXPHOS) activity, a prominent physiologic function of mitochondria. Cells lacking mitochondrial DNA or mutant cells with respiratory defects exhibited severely impaired virus-induced induction of interferons and proinflammatory cytokines. Recovery of the OXPHOS activity in these mutants, however, re-established RLR-mediated signal transduction. Using in vivo approaches, we found that mice with OXPHOS defects were highly susceptible to viral infection and exhibited significant lung inflammation. Studies to elucidate the molecular mechanism of OXPHOS-coupled immune activity revealed that optic atrophy 1, a mediator of mitochondrial fusion, contributes to regulate the antiviral immune response. Our findings provide evidence for functional coordination between RLR-mediated antiviral innate immunity and the mitochondrial energy-generating system in mammals. Introduction Innate immunity is a ubiquitous system that widely protects organisms from infectious pathogens as a front-line host defense mechanism. The immune response is triggered by the recognition of broadly conserved microbial components, known as pathogen-associated molecular patterns, by germline-encoded pattern recognition receptors of the host cells1. As an early defense system against RNA viruses in mammals, the innate immune response is precisely controlled by two distinct signal transduction pathways mediated by the pattern recognition receptors Toll-like receptor 3 (TLR-3) and retinoic acid-inducible gene I (RIG-I)-like receptors (RLR) that respond to virus-derived RNAs2, 3 (i.e., pathogen-associated molecular patterns). Although the two pathways differ with respect to the initial activation of their downstream effectors, they converge at the point of activation of the transcriptional factors interferon regulatory factor 3 (IRF-3) and nuclear factor κB (NF-κB), which results in the rapid production of type I interferons (IFN-α and -β) and other proinflammatory cytokines to establish adaptive antiviral immunity4. Mitochondria, eukaryotic cell powerhouses, are crucially involved in numerous cellular processes, including apoptosis5 and calcium homeostasis6. Mitochondria also have a unique role in innate immunity against RNA viruses7. Mitochondrial-mediated antiviral immunity depends on activation of the RLR signaling pathway, and mitochondrial antiviral signaling (MAVS), a downstream adaptor of RLR at the mitochondrial outer membrane (MOM), has a key role in the signal transduction8, 9. Upon viral infection, MAVS recruits various types of effectors at the MOM, and the orchestrated “MAVS signalosome”, including the mitochondrial membrane potential (Δψm), is the primary unit governing antiviral innate immunity10, 11. Although the role of the MAVS signalosome in mitochondria with its dynamic morphologic properties12 to provide a molecular platform that facilitates signal transduction is well characterized, insight into how the organelle functions to facilitate antiviral immunity through the activity of oxidative phosphorylation (OXPHOS) has remained unclear. Results and Discussion Cultured cells rely on mitochondrial respiratory activity To evaluate the functional coordination of mitochondrial-mediated antiviral immunity and OXPHOS activity, we first sought to determine optimal cell culture conditions in which the cellular bioenergetics would rely on mitochondrial respiratory activity. We used a fluorescence resonance energy transfer (FRET)-based assay to visualize metabolized intracellular adenosine 5′-triphosphate (ATP) at the single-cell level in human embryonic kidney 293 (HEK293) cells expressing an ATP probe, ATeam1.0313. The biosensor assay performed with cells cultured under our customized medium containing galactose (10 mM) as the carbon source revealed high FRET signal [based on an emission ratio of 527/475 nm (denoted YFP/CFP)] in individual cells [Fig. 1A, galactose panels, (−)], indicating that cells maintained adequate intracellular ATP levels. The intracellular ATP level, however, which affects the YFP/CFP ratio, was dramatically decreased (~2.5-fold) by the addition of electron transport chain (ETC) inhibitors (rotenone and antimycin A), an ATP synthase inhibitor (oligomycin), or a protonophore [carbonyl cyanide m-chlorophenylhydrazone (CCCP)] to the media [Fig. 1A, galactose panels, (+), and B], demonstrating that energy production in the cells critically depends on mitochondrial OXPHOS. In contrast, cells maintained in a customized medium containing glucose (10 mM) exhibited no significant decrease in the intracellular ATP level, even after treatment with the pharmacologic drugs ( (...truncated)


This is a preview of a remote PDF: https://www.nature.com/articles/s41598-017-05808-w.pdf

Takuma Yoshizumi, Hiromi Imamura, Tomohiro Taku, Takahiro Kuroki, Atsushi Kawaguchi, Kaori Ishikawa, Kazuto Nakada, Takumi Koshiba. RLR-mediated antiviral innate immunity requires oxidative phosphorylation activity, Scientific Reports, 2017, Issue: 7, DOI: 10.1038/s41598-017-05808-w