RESUMO
BACKGROUND & AIMS: Selective elimination of virus-infected hepatocytes occurs through virus-specific CD8 T cells recognizing peptide-loaded MHC molecules. Herein, we report that virus-infected hepatocytes are also selectively eliminated through a cell-autonomous mechanism. METHODS: We generated recombinant adenoviruses and genetically modified mouse models to identify the molecular mechanisms determining TNF-induced hepatocyte apoptosis in vivo and used in vivo bioluminescence imaging, immunohistochemistry, immunoblot analysis, RNAseq/proteome/phosphoproteome analyses, bioinformatic analyses, mitochondrial function tests. RESULTS: We found that TNF precisely eliminated only virus-infected hepatocytes independently of local inflammation and activation of immune sensory receptors. TNF receptor I was equally relevant for NF-kB activation in healthy and infected hepatocytes, but selectively mediated apoptosis in infected hepatocytes. Caspase 8 activation downstream of TNF receptor signaling was dispensable for apoptosis in virus-infected hepatocytes, indicating an unknown non-canonical cell-intrinsic pathway promoting apoptosis in hepatocytes. We identified a unique state of mitochondrial vulnerability in virus-infected hepatocytes as the cause for this non-canonical induction of apoptosis through TNF. Mitochondria from virus-infected hepatocytes showed normal biophysical and bioenergetic functions but were characterized by reduced resilience to calcium challenge. In the presence of unchanged TNF-induced signaling, reactive oxygen species-mediated calcium release from the endoplasmic reticulum caused mitochondrial permeability transition and apoptosis, which identified a link between extrinsic death receptor signaling and cell-intrinsic mitochondrial-mediated caspase activation. CONCLUSION: Our findings reveal a novel concept in immune surveillance by identifying a cell-autonomous defense mechanism that selectively eliminates virus-infected hepatocytes through mitochondrial permeability transition. LAY SUMMARY: The liver is known for its unique immune functions. Herein, we identify a novel mechanism by which virus-infected hepatocytes can selectively eliminate themselves through reduced mitochondrial resilience to calcium challenge.
Assuntos
Caspase 8/metabolismo , Hepatócitos , Mitocôndrias Hepáticas , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Animais , Apoptose/imunologia , Sinalização do Cálcio , Células Cultivadas , Hepatócitos/metabolismo , Hepatócitos/virologia , Humanos , Camundongos , Mitocôndrias Hepáticas/imunologia , Mitocôndrias Hepáticas/metabolismo , Necrose Dirigida por Permeabilidade Transmembrânica da Mitocôndria , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismoAssuntos
Infecções por Adenoviridae/imunologia , Infecções por Adenoviridae/metabolismo , Pneumonia Viral/imunologia , Pneumonia Viral/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Receptor Toll-Like 9/metabolismo , Adenoviridae , Infecções por Adenoviridae/virologia , Animais , Biomarcadores , Modelos Animais de Doenças , Camundongos , Pneumonia Viral/virologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismoRESUMO
Mitochondria are key for cellular metabolism and signalling processes during viral infection. We report a methodology to analyse mitochondrial properties at the single-organelle level during viral infection using a recombinant adenovirus coding for a mitochondrial tracer protein for tagging and detection by multispectral flow cytometry. Resolution at the level of tagged individual mitochondria revealed changes in mitochondrial size, membrane potential and displayed a fragile phenotype during viral infection of cells. Thus, single-organelle and multi-parameter resolution allows to explore altered energy metabolism and antiviral defence by tagged mitochondria selectively in virus-infected cells and will be instrumental to identify viral immune escape and to develop and monitor novel mitochondrial-targeted therapies.