Tetracycline-induced mitohormesis mediates disease tolerance against influenza
Adrienne Mottis, Terytty Y. Li, Gaby El Alam, Alexis Rapin, Elena Katsyuba, David Liaskos, Davide D’Amico, Nicola L. Harris, Mark C. Grier, Laurent Mouchiroud, Mark L. Nelson, Johan Auwerx
Adrienne Mottis, Terytty Y. Li, Gaby El Alam, Alexis Rapin, Elena Katsyuba, David Liaskos, Davide D’Amico, Nicola L. Harris, Mark C. Grier, Laurent Mouchiroud, Mark L. Nelson, Johan Auwerx
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Research Article Infectious disease

Tetracycline-induced mitohormesis mediates disease tolerance against influenza

Abstract

Mitohormesis defines the increase in fitness mediated by adaptive responses to mild mitochondrial stress. Tetracyclines inhibit not only bacterial but also mitochondrial translation, thus imposing a low level of mitochondrial stress on eukaryotic cells. We demonstrate in cell and germ-free mouse models that tetracyclines induce a mild adaptive mitochondrial stress response (MSR), involving both the ATF4-mediated integrative stress response and type I interferon (IFN) signaling. To overcome the interferences of tetracyclines with the host microbiome, we identify tetracycline derivatives that have minimal antimicrobial activity, yet retain full capacity to induce the MSR, such as the lead compound, 9-tert-butyl doxycycline (9-TB). The MSR induced by doxycycline (Dox) and 9-TB improves survival and disease tolerance against lethal influenza virus (IFV) infection when given preventively. 9-TB, unlike Dox, did not affect the gut microbiome and also showed encouraging results against IFV when given in a therapeutic setting. Tolerance to IFV infection is associated with the induction of genes involved in lung epithelial cell and cilia function, and with downregulation of inflammatory and immune gene sets in lungs, liver, and kidneys. Mitohormesis induced by non-antimicrobial tetracyclines and the ensuing IFN response may dampen excessive inflammation and tissue damage during viral infections, opening innovative therapeutic avenues.

Authors

Adrienne Mottis, Terytty Y. Li, Gaby El Alam, Alexis Rapin, Elena Katsyuba, David Liaskos, Davide D’Amico, Nicola L. Harris, Mark C. Grier, Laurent Mouchiroud, Mark L. Nelson, Johan Auwerx

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Figure 1

Doxycycline induces the ATF4 response and the type I IFN response.

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Doxycycline induces the ATF4 response and the type I IFN response. (A) B...
(A) Biochemical measurement of oxidative phosphorylation (OXPHOS) complexes (CI–CV), citrate synthase (CS), and ATP levels in the kidney of germ-free C57BL/6J male mice raised and maintained in a germ-free environment and that were drinking regular water or water supplemented with doxycycline (Dox) at 500 mg/kg/day (mpkd) for 16 days (n = 4–5). (B and C) Enrichment score plot for the gene set “Reactome Activation of genes by ATF4” (B) and heatmap representing the transcript levels of ATF4/5 targets (C) from kidney transcriptomics data of control versus Dox-treated germ-free mice. (D) Western blot analysis of selected ATF4 targets in the kidneys of germ-free mice (corresponding loading control below, HSP90). (E) Immunoblots of phosphorylated EIF2α (p-EIF2α) and total EIF2α in kidneys of Dox-treated germ-free mice. (F and G) Enrichment score plot for the GO term “Response to type I interferon” (F) and heatmap representing the transcript levels of some IFN-stimulated genes (ISGs) (G) from livers of germ-free mice treated with Dox. (H) Immunoblots of phosphorylated TBK1 (p-TBK1), TBK1, and the ISG proteins CGAS and CXCL10 (corresponding loading control below, vinculin and GAPDH, respectively). (I) Transcript levels of selected ISGs of bone marrow–derived macrophages (BMDMs) (day 6 of differentiation, derived from C57BL/6J mice) treated with Dox at 30 μg/mL for 9 hours (n = 6). (J) Immunoblots of phosphorylated TBK1 (p-TBK1), TBK1, and vinculin as control in BMDMs treated with Dox at 30 μg/mL for 3 hours. (K) Amplification of different mtDNA regions by qPCR in the cytosolic fraction of BMDMs with Dox at 30 μg/mL for 1 hour (n = 10). (L) Levels of IFN-β in the culture medium of BMDMs treated with Dox (30 μg/mL for 14 hours) and/or 2′,3′-dideoxycytidine (ddC, at 100 μM for 72 hours) (n = 8). Statistical analysis: Wilcoxon’s test P values corrected for multiple comparisons with Hommel’s method (A, I, and K) or by 1-way ANOVA followed by Tukey’s post hoc correction (L). *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001. NS, P > 0.05. Error bars represent ±SEM.