Abstract
Autophagy is a tightly regulated mechanism that mediates sequestration, degradation, and recycling of cellular proteins, organelles, and pathogens. Several proteins associated with autophagy regulate host responses to viral infections. Ribonuclease L (RNase L) is activated during viral infections and cleaves cellular and viral single-stranded RNAs, including rRNAs in ribosomes. Here we demonstrate that direct activation of RNase L coordinates the activation of c-Jun N-terminal kinase (JNK) and double-stranded RNA-dependent protein kinase (PKR) to induce autophagy with hallmarks as accumulation of autophagic vacuoles, p62(SQSTM1) degradation and conversion of Microtubule-associated Protein Light Chain 3-I (LC3-I) to LC3-II. Accordingly, treatment of cells with pharmacological inhibitors of JNK or PKR and mouse embryonic fibroblasts (MEFs) lacking JNK1/2 or PKR showed reduced autophagy levels. Furthermore, RNase L-induced JNK activity promoted Bcl-2 phosphorylation, disrupted the Beclin1-Bcl-2 complex and stimulated autophagy. Viral infection with Encephalomyocarditis virus (EMCV) or Sendai virus led to higher levels of autophagy in wild-type (WT) MEFs compared with RNase L knock out (KO) MEFs. Inhibition of RNase L-induced autophagy using Bafilomycin A1 or 3-methyladenine suppressed viral growth in initial stages; in later stages autophagy promoted viral replication dampening the antiviral effect. Induction of autophagy by activated RNase L is independent of the paracrine effects of interferon (IFN). Our findings suggest a novel role of RNase L in inducing autophagy affecting the outcomes of viral pathogenesis.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Adaptor Proteins, Signal Transducing / genetics
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Adaptor Proteins, Signal Transducing / metabolism
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Animals
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Autophagy*
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Cardiovirus Infections / genetics
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Cardiovirus Infections / metabolism
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Cardiovirus Infections / pathology
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Cell Line, Tumor
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Embryo, Mammalian / metabolism
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Embryo, Mammalian / pathology
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Encephalomyocarditis virus / physiology
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Endoribonucleases / genetics
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Endoribonucleases / metabolism*
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Fibroblasts / metabolism
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Fibroblasts / pathology
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Heat-Shock Proteins / genetics
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Heat-Shock Proteins / metabolism
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Humans
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MAP Kinase Signaling System*
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Mice
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Mice, Knockout
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Microtubule-Associated Proteins / genetics
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Microtubule-Associated Proteins / metabolism
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Mitogen-Activated Protein Kinase 8 / genetics
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Mitogen-Activated Protein Kinase 8 / metabolism*
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Mitogen-Activated Protein Kinase 9 / genetics
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Mitogen-Activated Protein Kinase 9 / metabolism*
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Proto-Oncogene Proteins c-bcl-2 / genetics
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Proto-Oncogene Proteins c-bcl-2 / metabolism
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RNA, Double-Stranded / genetics
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RNA, Double-Stranded / metabolism
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Respirovirus Infections / genetics
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Respirovirus Infections / metabolism
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Respirovirus Infections / pathology
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Sendai virus / physiology
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Sequestosome-1 Protein
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Virus Replication / physiology
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eIF-2 Kinase / genetics
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eIF-2 Kinase / metabolism*
Substances
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Adaptor Proteins, Signal Transducing
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Heat-Shock Proteins
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MAP1LC3A protein, human
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Map1lc3b protein, mouse
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Microtubule-Associated Proteins
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Proto-Oncogene Proteins c-bcl-2
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RNA, Double-Stranded
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SQSTM1 protein, human
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Sequestosome-1 Protein
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Sqstm1 protein, mouse
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Mitogen-Activated Protein Kinase 9
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eIF-2 Kinase
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Mitogen-Activated Protein Kinase 8
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Endoribonucleases
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2-5A-dependent ribonuclease