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Chen Z, Li Z, Wang Y, Dushimova Z, Gulnara K, Takeda S, Zhou Z, Xu X. ISGylation: is our genome yearning for such a modification? Acta Biochim Biophys Sin (Shanghai) 2025. [PMID: 40103488 DOI: 10.3724/abbs.2025028] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/20/2025] Open
Abstract
ISGylation is the post-translational modification of protein substrates covalently conjugated with the ubiquitin-like protein, interferon-stimulated gene 15 (ISG15). Although initially linked to antiviral immunity, recent evidence highlights important roles for ISGylation in various biological processes, such as maintaining genomic stability, promoting tumourigenesis, and being involved in other pathological conditions. In this review, we examine the molecular mechanisms underlying ISGylation, its interplay with other post-translational modifications, and its involvement in diverse biological and pathological processes. We propose future research directions to advance the field and discuss how ISGylation might be harnessed to ensure human health, particularly genome instability-associated diseases.
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Affiliation(s)
- Zheng Chen
- Shenzhen University General Hospital-Dehua Hospital Joint Research Center on Precision Medicine (sgh-dhhCPM), Dehua Hospital, Dehua 362500, China
- State Key Laboratory of Agrobiotechnology and MOA Key Laboratory of Soil Microbiology, College of Biological Sciences, China Agricultural University, Beijing 100193, China
- Guangdong Key Laboratory for Genome Stability & Disease Prevention and Carson International Cancer Center, Marshall Laboratory of Biomedical Engineering, Shenzhen University Medical School, Shenzhen 518060, China
| | - Zheng Li
- Shenzhen University General Hospital-Dehua Hospital Joint Research Center on Precision Medicine (sgh-dhhCPM), Dehua Hospital, Dehua 362500, China
- Guangdong Key Laboratory for Genome Stability & Disease Prevention and Carson International Cancer Center, Marshall Laboratory of Biomedical Engineering, Shenzhen University Medical School, Shenzhen 518060, China
| | - Ying Wang
- State Key Laboratory of Agrobiotechnology and MOA Key Laboratory of Soil Microbiology, College of Biological Sciences, China Agricultural University, Beijing 100193, China
| | - Zaure Dushimova
- Al-Farabi Kazakh National University, 71 Al-Farabi Avenue, Almaty 050040, Kazakhstan
| | - Kapanova Gulnara
- Al-Farabi Kazakh National University, 71 Al-Farabi Avenue, Almaty 050040, Kazakhstan
| | - Shunichi Takeda
- Guangdong Key Laboratory for Genome Stability & Disease Prevention and Carson International Cancer Center, Marshall Laboratory of Biomedical Engineering, Shenzhen University Medical School, Shenzhen 518060, China
| | - Zhongjun Zhou
- School of Biomedical Sciences, University of Hong Kong, Hong Kong 999077, China
| | - Xingzhi Xu
- Guangdong Key Laboratory for Genome Stability & Disease Prevention and Carson International Cancer Center, Marshall Laboratory of Biomedical Engineering, Shenzhen University Medical School, Shenzhen 518060, China
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2
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Bredow C, Thery F, Wirth EK, Ochs S, Kespohl M, Kleinau G, Kelm N, Gimber N, Schmoranzer J, Voss M, Klingel K, Spranger J, Renko K, Ralser M, Mülleder M, Heuser A, Knobeloch KP, Scheerer P, Kirwan J, Brüning U, Berndt N, Impens F, Beling A. ISG15 blocks cardiac glycolysis and ensures sufficient mitochondrial energy production during Coxsackievirus B3 infection. Cardiovasc Res 2024; 120:644-657. [PMID: 38309955 PMCID: PMC11074791 DOI: 10.1093/cvr/cvae026] [Citation(s) in RCA: 8] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/20/2023] [Revised: 11/10/2023] [Accepted: 12/12/2023] [Indexed: 02/05/2024] Open
Abstract
AIMS Virus infection triggers inflammation and, may impose nutrient shortage to the heart. Supported by type I interferon (IFN) signalling, cardiomyocytes counteract infection by various effector processes, with the IFN-stimulated gene of 15 kDa (ISG15) system being intensively regulated and protein modification with ISG15 protecting mice Coxsackievirus B3 (CVB3) infection. The underlying molecular aspects how the ISG15 system affects the functional properties of respective protein substrates in the heart are unknown. METHODS AND RESULTS Based on the protective properties due to protein ISGylation, we set out a study investigating CVB3-infected mice in depth and found cardiac atrophy with lower cardiac output in ISG15-/- mice. By mass spectrometry, we identified the protein targets of the ISG15 conjugation machinery in heart tissue and explored how ISGylation affects their function. The cardiac ISGylome showed a strong enrichment of ISGylation substrates within glycolytic metabolic processes. Two control enzymes of the glycolytic pathway, hexokinase 2 (HK2) and phosphofructokinase muscle form (PFK1), were identified as bona fide ISGylation targets during infection. In an integrative approach complemented with enzymatic functional testing and structural modelling, we demonstrate that protein ISGylation obstructs the activity of HK2 and PFK1. Seahorse-based investigation of glycolysis in cardiomyocytes revealed that, by conjugating proteins, the ISG15 system prevents the infection-/IFN-induced up-regulation of glycolysis. We complemented our analysis with proteomics-based advanced computational modelling of cardiac energy metabolism. Our calculations revealed an ISG15-dependent preservation of the metabolic capacity in cardiac tissue during CVB3 infection. Functional profiling of mitochondrial respiration in cardiomyocytes and mouse heart tissue by Seahorse technology showed an enhanced oxidative activity in cells with a competent ISG15 system. CONCLUSION Our study demonstrates that ISG15 controls critical nodes in cardiac metabolism. ISG15 reduces the glucose demand, supports higher ATP production capacity in the heart, despite nutrient shortage in infection, and counteracts cardiac atrophy and dysfunction.
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MESH Headings
- Animals
- Humans
- Male
- Coxsackievirus Infections/metabolism
- Coxsackievirus Infections/virology
- Coxsackievirus Infections/genetics
- Cytokines/genetics
- Cytokines/metabolism
- Disease Models, Animal
- Energy Metabolism
- Enterovirus B, Human/pathogenicity
- Enterovirus B, Human/metabolism
- Glycolysis
- Host-Pathogen Interactions
- Mice, Inbred C57BL
- Mice, Knockout
- Mitochondria, Heart/metabolism
- Mitochondria, Heart/pathology
- Myocytes, Cardiac/metabolism
- Myocytes, Cardiac/virology
- Myocytes, Cardiac/pathology
- Protein Processing, Post-Translational
- Signal Transduction
- Ubiquitins/metabolism
- Ubiquitins/genetics
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Affiliation(s)
- Clara Bredow
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Institute of Biochemistry, Charitéplatz 1, 10117 Berlin, Germany
| | - Fabien Thery
- Department of Biomolecular Medicine, Ghent University, Ghent, Belgium
- VIB-UGent Center for Medical Biotechnology, Ghent, Belgium
| | - Eva Katrin Wirth
- Deutsches Zentrum für Herz-Kreislauf-Forschung, partner site Berlin, Berlin, Germany
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Department of Endocrinology, Diabetes and Nutrition, Berlin, Germany
| | - Sarah Ochs
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Institute of Biochemistry, Charitéplatz 1, 10117 Berlin, Germany
| | - Meike Kespohl
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Institute of Biochemistry, Charitéplatz 1, 10117 Berlin, Germany
- Deutsches Zentrum für Herz-Kreislauf-Forschung, partner site Berlin, Berlin, Germany
| | - Gunnar Kleinau
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Institute of Medical Physics and Biophysics, Group Protein X-ray Crystallography and Signal Transduction, Charitéplatz 1, Berlin, Germany
| | - Nicolas Kelm
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Institute of Biochemistry, Charitéplatz 1, 10117 Berlin, Germany
| | - Niclas Gimber
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Advanced Medical Bioimaging Core Facility, Berlin, Germany
| | - Jan Schmoranzer
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Advanced Medical Bioimaging Core Facility, Berlin, Germany
| | - Martin Voss
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Institute of Biochemistry, Charitéplatz 1, 10117 Berlin, Germany
| | - Karin Klingel
- University of Tübingen, Cardiopathology, Institute for Pathology and Neuropathology, Tübingen, Germany
| | - Joachim Spranger
- Deutsches Zentrum für Herz-Kreislauf-Forschung, partner site Berlin, Berlin, Germany
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Department of Endocrinology, Diabetes and Nutrition, Berlin, Germany
| | - Kostja Renko
- German Federal Institute for Risk Assessment (BfR), German Centre for the Protection of Laboratory Animals (Bf3R), Berlin, Germany
| | - Markus Ralser
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Core Facility—High-Throughput Mass Spectrometry, Berlin, Germany
| | - Michael Mülleder
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Core Facility—High-Throughput Mass Spectrometry, Berlin, Germany
| | - Arnd Heuser
- Max-Delbrueck-Center (MDC) for Molecular Medicine, Animal Phenotyping Platform, Berlin, Germany
| | - Klaus-Peter Knobeloch
- University of Freiburg, Institute of Neuropathology, Freiburg, Germany
- CIBSS - Centre for Integrative Biological Signalling Studies, University of Freiburg, Freiburg, Germany
| | - Patrick Scheerer
- Deutsches Zentrum für Herz-Kreislauf-Forschung, partner site Berlin, Berlin, Germany
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Institute of Medical Physics and Biophysics, Group Protein X-ray Crystallography and Signal Transduction, Charitéplatz 1, Berlin, Germany
| | - Jennifer Kirwan
- Berlin Institute of Health at Charité Universitätsmedizin, Metabolomics, Charitéplatz 1 Berlin 10117, Germany
| | - Ulrike Brüning
- Berlin Institute of Health at Charité Universitätsmedizin, Metabolomics, Charitéplatz 1 Berlin 10117, Germany
| | - Nikolaus Berndt
- German Institute of Human Nutrition Potsdam-Rehbruecke (DIfE), Department of Molecular Toxicology, Nuthetal, Germany
- Deutsches Herzzentrum der Charité (DHZC), Institute of Computer-assisted Cardiovascular Medicine, Berlin, Germany
- Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Berlin, Germany
| | - Francis Impens
- Department of Biomolecular Medicine, Ghent University, Ghent, Belgium
- VIB-UGent Center for Medical Biotechnology, Ghent, Belgium
- VIB Proteomics Core, Ghent, Belgium
| | - Antje Beling
- Charité - Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt-Universität zu Berlin, Institute of Biochemistry, Charitéplatz 1, 10117 Berlin, Germany
- Deutsches Zentrum für Herz-Kreislauf-Forschung, partner site Berlin, Berlin, Germany
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3
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Jové V, Wheeler H, Lee CW, Healy DR, Levine K, Ralph EC, Yamaguchi M, Jiang ZK, Cabral E, Xu Y, Stock J, Yang B, Giddabasappa A, Loria P, Casimiro-Garcia A, Kessler BM, Pinto-Fernández A, Frattini V, Wes PD, Wang F. Type I interferon regulation by USP18 is a key vulnerability in cancer. iScience 2024; 27:109593. [PMID: 38632987 PMCID: PMC11022047 DOI: 10.1016/j.isci.2024.109593] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/26/2023] [Revised: 01/12/2024] [Accepted: 03/25/2024] [Indexed: 04/19/2024] Open
Abstract
Precise regulation of Type I interferon signaling is crucial for combating infection and cancer while avoiding autoimmunity. Type I interferon signaling is negatively regulated by USP18. USP18 cleaves ISG15, an interferon-induced ubiquitin-like modification, via its canonical catalytic function, and inhibits Type I interferon receptor activity through its scaffold role. USP18 loss-of-function dramatically impacts immune regulation, pathogen susceptibility, and tumor growth. However, prior studies have reached conflicting conclusions regarding the relative importance of catalytic versus scaffold function. Here, we develop biochemical and cellular methods to systematically define the physiological role of USP18. By comparing a patient-derived mutation impairing scaffold function (I60N) to a mutation disrupting catalytic activity (C64S), we demonstrate that scaffold function is critical for cancer cell vulnerability to Type I interferon. Surprisingly, we discovered that human USP18 exhibits minimal catalytic activity, in stark contrast to mouse USP18. These findings resolve human USP18's mechanism-of-action and enable USP18-targeted therapeutics.
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Affiliation(s)
- Veronica Jové
- Centers for Therapeutic Innovation, Pfizer, New York City, NY 10016, USA
| | - Heather Wheeler
- Discovery Sciences, Medicine Design, Pfizer, Groton, CT 06340, USA
| | | | - David R. Healy
- Discovery Sciences, Medicine Design, Pfizer, Groton, CT 06340, USA
| | - Kymberly Levine
- Centers for Therapeutic Innovation, Pfizer, New York City, NY 10016, USA
| | - Erik C. Ralph
- Discovery Sciences, Medicine Design, Pfizer, Groton, CT 06340, USA
| | - Masaya Yamaguchi
- Discovery Sciences, Medicine Design, Pfizer, Groton, CT 06340, USA
| | | | - Edward Cabral
- Comparative Medicine, Pfizer, La Jolla, CA 92121, USA
| | - Yingrong Xu
- Discovery Sciences, Medicine Design, Pfizer, Groton, CT 06340, USA
| | - Jeffrey Stock
- Discovery Sciences, Medicine Design, Pfizer, Groton, CT 06340, USA
| | - Bing Yang
- Comparative Medicine, Pfizer, La Jolla, CA 92121, USA
| | | | - Paula Loria
- Discovery Sciences, Medicine Design, Pfizer, Groton, CT 06340, USA
| | | | - Benedikt M. Kessler
- Chinese Academy for Medical Sciences Oxford Institute, Nuffield Department of Medicine, University of Oxford, Oxford OX3 7BN, UK
- Target Discovery Institute, Centre for Medicines Discovery, Nuffield Department of Medicine, University of Oxford, Oxford OX3 7FZ, UK
| | - Adán Pinto-Fernández
- Chinese Academy for Medical Sciences Oxford Institute, Nuffield Department of Medicine, University of Oxford, Oxford OX3 7BN, UK
- Target Discovery Institute, Centre for Medicines Discovery, Nuffield Department of Medicine, University of Oxford, Oxford OX3 7FZ, UK
| | - Véronique Frattini
- Centers for Therapeutic Innovation, Pfizer, New York City, NY 10016, USA
| | - Paul D. Wes
- Centers for Therapeutic Innovation, Pfizer, New York City, NY 10016, USA
| | - Feng Wang
- Discovery Sciences, Medicine Design, Pfizer, Groton, CT 06340, USA
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4
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Xu Q, Li W, Zhao Q, Zhao L, Lv G, Sun G, Gao Y, Ding Y, Zhang Z, Zhou L, Chen Y, Tang X, Zhu J, Zhao X, An Y. A novel homozygous Y140X mutation of ISG15 causes diverse type I interferonopathies in sibling patients with cutaneous lesions or recurrent parenchymal pneumonia. Clin Immunol 2023; 257:109844. [PMID: 37984483 DOI: 10.1016/j.clim.2023.109844] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2023] [Revised: 10/25/2023] [Accepted: 11/13/2023] [Indexed: 11/22/2023]
Abstract
PURPOSE Interferon-stimulated gene 15 (ISG15) deficiency, a rare human inborn error of immunity characterized by susceptibility to Bacillus Calmette-Guerin (BCG) diseases, neuropathic and dermatological manifestations. METHODS The clinical and immunological features of two siblings with ISG15 deficiency combined with asymptomatic myeloperoxidase (MPO) mutations were analyzed, and their pathogenesis, as well as target therapeutic candidates, were explored. RESULTS The manifestation in patient 2 was skin lesions, while those in patient 1 were intracranial calcification and recurrent pneumonia. Whole-exome identified novel, dual mutations in ISG15 and MPO. PBMCs and B cell lines derived from the patients showed hyper-activated JAK/STAT signaling. Normal neutrophil function excluded pathogenicity caused by the MPO mutation. RNA sequencing identified baricitinib as therapeutic candidate. CONCLUSIONS We report two sibling patients harboring the same novel ISG15 mutation showing diverse clinical features, and one harbored a rare phenotype of pneumonia. These findings expand the clinical spectrum of ISG15 deficiency and identify baricitinib as therapeutic candidate.
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Affiliation(s)
- Qiling Xu
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Wenyan Li
- Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Qian Zhao
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Lu Zhao
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China
| | - Ge Lv
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Gan Sun
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Yelei Gao
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Yuan Ding
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Zhiyong Zhang
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China; Department of Rheumatology & Immunology, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Lina Zhou
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Yongwen Chen
- Institute of Immunology, PLA, Third Military Medical University, Chongqing 400014, People's Republic of China
| | - Xuemei Tang
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China; Department of Rheumatology & Immunology, Children's Hospital of Chongqing Medical University, Chongqing, China
| | - Jin Zhu
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Department of Pathology, College of Basic Medicine, Molecular 6. Medicine Diagnostic and Testing Center, Chongqing Medical University, Chongqing, China; Institute of Immunology, PLA, Third Military Medical University, Chongqing 400014, People's Republic of China
| | - Xiaodong Zhao
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China.
| | - Yunfei An
- Childrens Hospital of Chongqing Medical University, National Clinical Research Center for Child Health and Disorders (Chongqing), Chongqing, China; Ministry of Education Key Laboratory of Child Development and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Chongqing Key Laboratory of Child Infection and Immunity, Children's Hospital of Chongqing Medical University, Chongqing, China; Department of Rheumatology & Immunology, Children's Hospital of Chongqing Medical University, Chongqing, China.
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5
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Sarkar R, Patra U, Mukherjee A, Mitra S, Komoto S, Chawla-Sarkar M. Rotavirus circumvents the antiviral effects of protein ISGylation via proteasomal degradation of Ube1L. Cell Signal 2023; 112:110891. [PMID: 37722521 DOI: 10.1016/j.cellsig.2023.110891] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2023] [Revised: 08/10/2023] [Accepted: 09/15/2023] [Indexed: 09/20/2023]
Abstract
Among the ramified cellular responses elicited in response to pathogenic stimuli, upregulation and covalent conjugation of an Ubiquitin-like modifier ISG15 to lysine residues of target proteins (ISGylation) through sequential action of three enzymes E1 (Ube1L), E2 (Ube2L6) and E3 (Herc5) have emerged as an important regulatory facet governing innate immunity against numerous viral infections. In the present study, we investigated the interplay between host ISGylation system and Rotavirus (RV). We observed that RV infection upregulates the expression of free ISG15 but prevents protein ISGylation. Analysing the expression of ISGylation machinery components revealed that RV infection results in steady depletion of Ube1L protein with the progression of infection. Indeed, restoration of Ube1L expression caused induction in protein ISGylation during RV infection. Subsequent investigation revealed that ectopic expression of RV non-structural protein 5 (NSP5) fosters proteolytic ubiquitylation of Ube1L, thereby depleting it in an ubiquitin-proteasome-dependent manner. Moreover, pan-Cullin inhibition also abrogates proteolytic ubiquitylation and rescued depleted Ube1L in RV-NSP5 expressing cells, suggesting the involvement of host cellular Cullin RING Ligases (CRLs) in proteasomal degradation of Ube1L during RV-SA11 infection. Reciprocal co-immunoprecipitation analyses substantiated a molecular association between Ube1L and RV-NSP5 during infection scenario and also under ectopically overexpressed condition independent of intermediate RNA scaffold and RV-NSP5 hyperphosphorylation. Interestingly, clonal overexpression of Ube1L reduced expression of RV proteins and RV infectivity, which are restored in ISG15 silenced cells, suggesting that Ube1L is a crucial anti-viral host cellular determinant that inhibits RV infection by promoting the formation of ISG15 conjugates.
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Affiliation(s)
- Rakesh Sarkar
- Division of Virology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, West Bengal, India
| | - Upayan Patra
- Division of Virology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, West Bengal, India
| | - Arpita Mukherjee
- Division of Virology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, West Bengal, India
| | - Suvrotoa Mitra
- Division of Virology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, West Bengal, India
| | - Satoshi Komoto
- Department of Virology and Parasitology, School of Medicine, Fujita Health University, Aichi, Japan
| | - Mamta Chawla-Sarkar
- Division of Virology, ICMR-National Institute of Cholera and Enteric Diseases, Kolkata, West Bengal, India.
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6
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Si Y, Zhang H, Zhou Z, Zhu X, Yang Y, Liu H, Zhang L, Cheng L, Wang K, Ye W, Lv X, Zhang X, Hou W, Zhao G, Lei Y, Zhang F, Ma H. RIPK3 promotes hantaviral replication by restricting JAK-STAT signaling without triggering necroptosis. Virol Sin 2023; 38:741-754. [PMID: 37633447 PMCID: PMC10590702 DOI: 10.1016/j.virs.2023.08.006] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2023] [Accepted: 08/21/2023] [Indexed: 08/28/2023] Open
Abstract
Hantaan virus (HTNV) is a rodent-borne virus that causes hemorrhagic fever with renal syndrome (HFRS), resulting in a high mortality rate of 15%. Interferons (IFNs) play a critical role in the anti-hantaviral immune response, and IFN pretreatment efficiently restricts HTNV infection by triggering the expression of a series of IFN-stimulated genes (ISGs) through the Janus kinase-signal transducer and activator of transcription 1 (JAK-STAT) pathway. However, the tremendous amount of IFNs produced during late infection could not restrain HTNV replication, and the mechanism remains unclear. Here, we demonstrated that receptor-interacting protein kinase 3 (RIPK3), a crucial molecule that mediates necroptosis, was activated by HTNV and contributed to hantavirus evasion of IFN responses by inhibiting STAT1 phosphorylation. RNA-seq analysis revealed the upregulation of multiple cell death-related genes after HTNV infection, with RIPK3 identified as a key modulator of viral replication. RIPK3 ablation significantly enhanced ISGs expression and restrained HTNV replication, without affecting the expression of pattern recognition receptors (PRRs) or the production of type I IFNs. Conversely, exogenously expressed RIPK3 compromised the host's antiviral response and facilitated HTNV replication. RIPK3-/- mice also maintained a robust ability to clear HTNV with enhanced innate immune responses. Mechanistically, we found that RIPK3 could bind STAT1 and inhibit STAT1 phosphorylation dependent on the protein kinase domain (PKD) of RIPK3 but not its kinase activity. Overall, these observations demonstrated a noncanonical function of RIPK3 during viral infection and have elucidated a novel host innate immunity evasion strategy utilized by HTNV.
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Affiliation(s)
- Yue Si
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China
| | - Haijun Zhang
- Department of Neurology, Xijing Hospital, Air Force Medical University, Xi'an, 710032, China; Center of Clinical Aerospace Medicine, School of Aerospace Medicine, Key Laboratory of Aerospace Medicine of Ministry of Education, Air Force Medical University, Xi'an, 710032, China
| | - Ziqing Zhou
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China
| | - Xudong Zhu
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China
| | - Yongheng Yang
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China
| | - He Liu
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China
| | - Liang Zhang
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China
| | - Linfeng Cheng
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China
| | - Kerong Wang
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China
| | - Wei Ye
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China
| | - Xin Lv
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China
| | - Xijing Zhang
- Department of Anesthesiology & Critical Care Medicine, Xijing Hospital, Air Force Medical University, Xi'an, 710032, China
| | - Wugang Hou
- Department of Anesthesiology & Critical Care Medicine, Xijing Hospital, Air Force Medical University, Xi'an, 710032, China
| | - Gang Zhao
- Department of Neurology, Xijing Hospital, Air Force Medical University, Xi'an, 710032, China; The College of Life Sciences and Medicine, Northwest University, Xi'an, 710069, China
| | - Yingfeng Lei
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China.
| | - Fanglin Zhang
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China.
| | - Hongwei Ma
- Department of Microbiology, School of Basic Medicine, Air Force Medical University, Xi'an, 710032, China; Department of Anesthesiology & Critical Care Medicine, Xijing Hospital, Air Force Medical University, Xi'an, 710032, China.
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7
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Li L, Miao J, Shaheen N, Taleb SJ, Hu J, Ye Q, He J, Yan J, Mallampalli RK, Zhao J, Zhao Y. ISGylation of NF-κBp65 by SCF FBXL19 E3 Ligase Diminishes Endothelial Inflammation. Arterioscler Thromb Vasc Biol 2023; 43:674-683. [PMID: 36994728 PMCID: PMC10133096 DOI: 10.1161/atvbaha.122.318894] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2022] [Accepted: 03/21/2023] [Indexed: 03/31/2023]
Abstract
BACKGROUND NF-κB (nuclear factor kappa B) plays a pivotal role in endothelial cell (EC) inflammation. Protein ISGylation is regulated by E3 ISG15 (interferon-stimulated gene 15) ligases; however, ISGylation of NF-κBp65 and its role in EC functions have not been investigated. Here, we investigate whether p65 is ISGylated and the role of its ISGylation in endothelial functions. METHODS In vitro ISGylation assay and EC inflammation were performed. EC-specific transgenic mice were utilized in a murine model of acute lung injury. RESULTS We find that NF-κBp65 is ISGylated in resting ECs and that the posttranslational modification is reversible. TNFα (tumor necrosis factor alpha) and endotoxin stimulation of EC reduce p65 ISGylation, promoting its serine phosphorylation through reducing its association with a phosphatase WIP1 (wild-type p53-induced phosphatase 1). Mechanistically, an SCF (Skp1-Cul1-F-box) protein E3 ligase SCFFBXL19 is identified as a new ISG15 E3 ligase that targets and catalyzes ISGylation of p65. Depletion of FBXL19 (F-box and leucine-rich repeat protein 19) increases p65 phosphorylation and EC inflammation, suggesting a negative correlation between p65 ISGylation and phosphorylation. Moreover, EC-specific FBXL19 overexpressing humanized transgenic mice exhibit reduced lung inflammation and severity of experimental acute lung injury. CONCLUSIONS Together, our data reveal a new posttranslational modification of p65 catalyzed by a previously unrecognized role of SCFFBXL19 as an ISG15 E3 ligase that modulates EC inflammation.
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Affiliation(s)
- Lian Li
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart and Lung Research Institute, the Ohio State University, Columbus, OH
| | - Jiaxing Miao
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart and Lung Research Institute, the Ohio State University, Columbus, OH
| | - Nargis Shaheen
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart and Lung Research Institute, the Ohio State University, Columbus, OH
| | - Sarah J. Taleb
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart and Lung Research Institute, the Ohio State University, Columbus, OH
| | - Jian Hu
- Department of Internal Medicine, the Ohio State University, Columbus, OH
| | - Qinmao Ye
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart and Lung Research Institute, the Ohio State University, Columbus, OH
| | - Jinshan He
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart and Lung Research Institute, the Ohio State University, Columbus, OH
| | - Jiasheng Yan
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart and Lung Research Institute, the Ohio State University, Columbus, OH
| | | | - Jing Zhao
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart and Lung Research Institute, the Ohio State University, Columbus, OH
- Department of Internal Medicine, the Ohio State University, Columbus, OH
| | - Yutong Zhao
- Department of Physiology and Cell Biology, Dorothy M. Davis Heart and Lung Research Institute, the Ohio State University, Columbus, OH
- Department of Internal Medicine, the Ohio State University, Columbus, OH
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8
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Huang L, Cheng Y, Han S, Liu M, Yu Q, Wei H, He J, Li P. Identification of ISG15 in golden pompano, Trachinotus ovatus, and its role in virus and bacteria infections. FISH & SHELLFISH IMMUNOLOGY 2023; 132:108481. [PMID: 36566833 DOI: 10.1016/j.fsi.2022.108481] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/10/2022] [Revised: 11/25/2022] [Accepted: 12/06/2022] [Indexed: 06/17/2023]
Abstract
Interferon (IFN)-stimulated gene product 15 (ISG15) is a ubiquitin-like protein critical for the control of microbial infections. Golden pompano, Trachinotus ovatus is one of the precious marine economic fish in the southern coast of China, always suffering from viruses, bacteria, and parasite infections. To date, the roles of golden pompano genes involved in viral and bacterial infections, especially IFN-related genes remained largely unknown. To identify the interferon system genes of golden pompano and explore their function, in this study, the ISG15 homolog (ToISG15) was cloned from golden pompano, and its role in response to grouper iridovirus (SGIV), nervous necrosis virus (NNV), and Aeromonas hydrophila infection was investigated. The whole ORF of ToISG15 was composed of 465 bp and encoded a polypeptide of 154 amino acids with different identity with the known ISG15 homologs from other fish species. Two conserved ubiquitin-like (UBL) domains and an Ub-conjugation domain (LRGG) were found in ToISG15 sequence. Expression analysis showed that ToISG15 was located mainly in the cytoplasm of golden pompano cells, and dramatically induced following SGIV, Aeromonas hydrophila, or poly I:C treatment, but little change was observed when NNV infection. Overexpression of ToISG15 in vitro significantly inhibited the replication of SGIV and NNV. Interestingly, ToISG15 possessed the ability to restrain the growth of Aeromonas hydrophila. Furthermore, To-ISG15 overexpression enhanced the expression of IFNc, IFNh, IRF3, IRF7, and viperin genes as well as, to a lesser extent, the IL-6 gene. Taken together, our results demonstrated the antiviral and antibacterial effect of To-ISG15, shedding light on the evolutionary conservation of ISG15 in the immune response to microbial infection.
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Affiliation(s)
- Lin Huang
- Guangxi Key Laboratory of Aquatic Biotechnology and Modern Ecological Aquaculture, Guangxi Engineering Research Center for Fishery Major Diseases Control and Efficient Healthy Breeding Industrial Technology (GERCFT), Guangxi Academy of Sciences, Nanning, China; China-ASEAN Modern Fishery Industry Technology Transfer Demonstration Center, Guangxi Academy of Sciences, Nanning, China
| | - Yuan Cheng
- Guangxi Key Laboratory of Aquatic Biotechnology and Modern Ecological Aquaculture, Guangxi Engineering Research Center for Fishery Major Diseases Control and Efficient Healthy Breeding Industrial Technology (GERCFT), Guangxi Academy of Sciences, Nanning, China
| | - Shuyu Han
- Guangxi Fisheries Technology Extension Station, Nanning, China
| | - Mingzhu Liu
- Guangxi Key Laboratory of Aquatic Biotechnology and Modern Ecological Aquaculture, Guangxi Engineering Research Center for Fishery Major Diseases Control and Efficient Healthy Breeding Industrial Technology (GERCFT), Guangxi Academy of Sciences, Nanning, China; China-ASEAN Modern Fishery Industry Technology Transfer Demonstration Center, Guangxi Academy of Sciences, Nanning, China
| | - Qing Yu
- Guangxi Key Laboratory of Aquatic Biotechnology and Modern Ecological Aquaculture, Guangxi Engineering Research Center for Fishery Major Diseases Control and Efficient Healthy Breeding Industrial Technology (GERCFT), Guangxi Academy of Sciences, Nanning, China; China-ASEAN Modern Fishery Industry Technology Transfer Demonstration Center, Guangxi Academy of Sciences, Nanning, China
| | - Hongling Wei
- Guangxi Key Laboratory of Aquatic Biotechnology and Modern Ecological Aquaculture, Guangxi Engineering Research Center for Fishery Major Diseases Control and Efficient Healthy Breeding Industrial Technology (GERCFT), Guangxi Academy of Sciences, Nanning, China; China-ASEAN Modern Fishery Industry Technology Transfer Demonstration Center, Guangxi Academy of Sciences, Nanning, China
| | - Jinzhao He
- Guangxi Fisheries Technology Extension Station, Nanning, China.
| | - Pengfei Li
- Guangxi Key Laboratory of Aquatic Biotechnology and Modern Ecological Aquaculture, Guangxi Engineering Research Center for Fishery Major Diseases Control and Efficient Healthy Breeding Industrial Technology (GERCFT), Guangxi Academy of Sciences, Nanning, China; China-ASEAN Modern Fishery Industry Technology Transfer Demonstration Center, Guangxi Academy of Sciences, Nanning, China.
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9
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Coronaviral PLpro proteases and the immunomodulatory roles of conjugated versus free Interferon Stimulated Gene product-15 (ISG15). Semin Cell Dev Biol 2022; 132:16-26. [PMID: 35764457 PMCID: PMC9233553 DOI: 10.1016/j.semcdb.2022.06.005] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/10/2021] [Revised: 06/12/2022] [Accepted: 06/13/2022] [Indexed: 12/14/2022]
Abstract
Ubiquitin-like proteins (Ubls) share some features with ubiquitin (Ub) such as their globular 3D structure and the ability to attach covalently to other proteins. Interferon Stimulated Gene 15 (ISG15) is an abundant Ubl that similar to Ub, marks many hundreds of cellular proteins, altering their fate. In contrast to Ub, , ISG15 requires interferon (IFN) induction to conjugate efficiently to other proteins. Moreover, despite the multitude of E3 ligases for Ub-modified targets, a single E3 ligase termed HERC5 (in humans) is responsible for the bulk of ISG15 conjugation. Targets include both viral and cellular proteins spanning an array of cellular compartments and metabolic pathways. So far, no common structural or biochemical feature has been attributed to these diverse substrates, raising questions about how and why they are selected. Conjugation of ISG15 mitigates some viral and bacterial infections and is linked to a lower viral load pointing to the role of ISG15 in the cellular immune response. In an apparent attempt to evade the immune response, some viruses try to interfere with the ISG15 pathway. For example, deconjugation of ISG15 appears to be an approach taken by coronaviruses to interfere with ISG15 conjugates. Specifically, coronaviruses such as SARS-CoV, MERS-CoV, and SARS-CoV-2, encode papain-like proteases (PL1pro) that bear striking structural and catalytic similarities to the catalytic core domain of eukaryotic deubiquitinating enzymes of the Ubiquitin-Specific Protease (USP) sub-family. The cleavage specificity of these PLpro enzymes is for flexible polypeptides containing a consensus sequence (R/K)LXGG, enabling them to function on two seemingly unrelated categories of substrates: (i) the viral polyprotein 1 (PP1a, PP1ab) and (ii) Ub- or ISG15-conjugates. As a result, PLpro enzymes process the viral polyprotein 1 into an array of functional proteins for viral replication (termed non-structural proteins; NSPs), and it can remove Ub or ISG15 units from conjugates. However, by de-conjugating ISG15, the virus also creates free ISG15, which in turn may affect the immune response in two opposite pathways: free ISG15 negatively regulates IFN signaling in humans by binding non-catalytically to USP18, yet at the same time free ISG15 can be secreted from the cell and induce the IFN pathway of the neighboring cells. A deeper understanding of this protein-modification pathway and the mechanisms of the enzymes that counteract it will bring about effective clinical strategies related to viral and bacterial infections.
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10
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Targeting papain-like protease for broad-spectrum coronavirus inhibition. Protein Cell 2022; 13:940-953. [PMID: 35384604 PMCID: PMC8983325 DOI: 10.1007/s13238-022-00909-3] [Citation(s) in RCA: 34] [Impact Index Per Article: 11.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2021] [Accepted: 02/17/2022] [Indexed: 11/25/2022] Open
Abstract
The emergence of SARS-CoV-2 variants of concern and repeated outbreaks of coronavirus epidemics in the past two decades emphasize the need for next-generation pan-coronaviral therapeutics. Drugging the multi-functional papain-like protease (PLpro) domain of the viral nsp3 holds promise. However, none of the known coronavirus PLpro inhibitors has been shown to be in vivo active. Herein, we screened a structurally diverse library of 50,080 compounds for potential coronavirus PLpro inhibitors and identified a noncovalent lead inhibitor F0213 that has broad-spectrum anti-coronaviral activity, including against the Sarbecoviruses (SARS-CoV-1 and SARS-CoV-2), Merbecovirus (MERS-CoV), as well as the Alphacoronavirus (hCoV-229E and hCoV-OC43). Importantly, F0213 confers protection in both SARS-CoV-2-infected hamsters and MERS-CoV-infected human DPP4-knockin mice. F0213 possesses a dual therapeutic functionality that suppresses coronavirus replication via blocking viral polyprotein cleavage, as well as promoting antiviral immunity by antagonizing the PLpro deubiquitinase activity. Despite the significant difference of substrate recognition, mode of inhibition studies suggest that F0213 is a competitive inhibitor against SARS2-PLpro via binding with the 157K amino acid residue, whereas an allosteric inhibitor of MERS-PLpro interacting with its 271E position. Our proof-of-concept findings demonstrated that PLpro is a valid target for the development of broad-spectrum anti-coronavirus agents. The orally administered F0213 may serve as a promising lead compound for combating the ongoing COVID-19 pandemic and future coronavirus outbreaks.
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11
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Xiong TC, Wei MC, Li FX, Shi M, Gan H, Tang Z, Dong HP, Liuyu T, Gao P, Zhong B, Zhang ZD, Lin D. The E3 ubiquitin ligase ARIH1 promotes antiviral immunity and autoimmunity by inducing mono-ISGylation and oligomerization of cGAS. Nat Commun 2022; 13:5973. [PMID: 36217001 PMCID: PMC9551088 DOI: 10.1038/s41467-022-33671-5] [Citation(s) in RCA: 39] [Impact Index Per Article: 13.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/18/2022] [Accepted: 09/26/2022] [Indexed: 11/09/2022] Open
Abstract
The cytosolic DNA sensor cyclic GMP-AMP synthase (cGAS) plays a critical role in antiviral immunity and autoimmunity. The activity and stability of cGAS are fine-tuned by post-translational modifications. Here, we show that ariadne RBR E3 ubiquitin protein ligase 1 (ARIH1) catalyzes the mono-ISGylation and induces the oligomerization of cGAS, thereby promoting antiviral immunity and autoimmunity. Knockdown or knockout of ARIH1 significantly inhibits herpes simplex virus 1 (HSV-1)- or cytoplasmic DNA-induced expression of type I interferons (IFNs) and proinflammatory cytokines. Consistently, tamoxifen-treated ER-Cre;Arih1fl/fl mice and Lyz2-Cre; Arih1fl/fl mice are hypersensitive to HSV-1 infection compared with the controls. In addition, deletion of ARIH1 in myeloid cells alleviates the autoimmune phenotypes and completely rescues the autoimmune lethality caused by TREX1 deficiency. Mechanistically, HSV-1- or cytosolic DNA-induced oligomerization and activation of cGAS are potentiated by ISGylation at its K187 residue, which is catalyzed by ARIH1. Our findings thus reveal an important role of ARIH1 in innate antiviral and autoimmune responses and provide insight into the post-translational regulation of cGAS.
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Affiliation(s)
- Tian-Chen Xiong
- Department of Gastrointestinal Surgery, Medical Research Institute, Frontier Science Center of Immunology and Metabolism, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China
- Chongqing International Institute for Immunology, Chongqing, China
- TaiKang Center for Life and Medical Sciences, Wuhan University, Wuhan, China
- Wuhan Research Center for Infectious Diseases and Cancer, Chinese Academy of Medical Sciences, Wuhan, China
| | - Ming-Cong Wei
- Department of Gastrointestinal Surgery, Medical Research Institute, Frontier Science Center of Immunology and Metabolism, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China
- TaiKang Center for Life and Medical Sciences, Wuhan University, Wuhan, China
| | - Fang-Xu Li
- Department of Gastrointestinal Surgery, Medical Research Institute, Frontier Science Center of Immunology and Metabolism, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China
- TaiKang Center for Life and Medical Sciences, Wuhan University, Wuhan, China
| | - Miao Shi
- CAS Key Laboratory of Infection and Immunity, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China
| | - Hu Gan
- Department of Gastrointestinal Surgery, Medical Research Institute, Frontier Science Center of Immunology and Metabolism, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China
- TaiKang Center for Life and Medical Sciences, Wuhan University, Wuhan, China
- Department of Virology, College of Life Sciences, Wuhan University, Wuhan, China
| | - Zhen Tang
- Department of Gastrointestinal Surgery, Medical Research Institute, Frontier Science Center of Immunology and Metabolism, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China
- TaiKang Center for Life and Medical Sciences, Wuhan University, Wuhan, China
- Department of Virology, College of Life Sciences, Wuhan University, Wuhan, China
| | - Hong-Peng Dong
- Department of Gastrointestinal Surgery, Medical Research Institute, Frontier Science Center of Immunology and Metabolism, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China
- TaiKang Center for Life and Medical Sciences, Wuhan University, Wuhan, China
| | - Tianzi Liuyu
- Department of Gastrointestinal Surgery, Medical Research Institute, Frontier Science Center of Immunology and Metabolism, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China
- TaiKang Center for Life and Medical Sciences, Wuhan University, Wuhan, China
| | - Pu Gao
- CAS Key Laboratory of Infection and Immunity, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China
| | - Bo Zhong
- Department of Gastrointestinal Surgery, Medical Research Institute, Frontier Science Center of Immunology and Metabolism, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China.
- TaiKang Center for Life and Medical Sciences, Wuhan University, Wuhan, China.
- Wuhan Research Center for Infectious Diseases and Cancer, Chinese Academy of Medical Sciences, Wuhan, China.
- Department of Virology, College of Life Sciences, Wuhan University, Wuhan, China.
| | - Zhi-Dong Zhang
- Department of Gastrointestinal Surgery, Medical Research Institute, Frontier Science Center of Immunology and Metabolism, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan, China.
| | - Dandan Lin
- Cancer Center, Renmin Hospital of Wuhan University, Wuhan, China.
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12
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Liu H, Li C, He W, Chen J, Yang G, Chen L, Chang H. Free ISG15 inhibits Pseudorabies virus infection by positively regulating type I IFN signaling. PLoS Pathog 2022; 18:e1010921. [PMID: 36315588 PMCID: PMC9648840 DOI: 10.1371/journal.ppat.1010921] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2022] [Revised: 11/10/2022] [Accepted: 10/07/2022] [Indexed: 11/12/2022] Open
Abstract
Interferon-stimulated gene 15 (ISG15) is strongly upregulated during viral infections and exerts pro-viral or antiviral actions. While many viruses combat host antiviral defenses by limiting ISG expression, PRV infection notably increases expression of ISG15. However, studies on the viral strategies to regulate ISG15-mediated antiviral responses are limited. Here, we demonstrate that PRV-induced free ISG15 and conjugated proteins accumulation require viral gene expression. Conjugation inhibition assays showed that ISG15 imposes its antiviral effects via unconjugated (free) ISG15 and restricts the viral release. Knockout of ISG15 in PK15 cells interferes with IFN-β production by blocking IRF3 activation and promotes PRV replication. Mechanistically, ISG15 facilitates IFNα-mediated antiviral activity against PRV by accelerating the activation and nuclear translocation of STAT1 and STAT2. Furthermore, ISG15 facilitated STAT1/STAT2/IRF9 (ISGF3) formation and ISGF3-induced IFN-stimulated response elements (ISRE) activity for efficient gene transcription by directly interacting with STAT2. Significantly, ISG15 knockout mice displayed enhanced susceptibility to PRV, as evidenced by increased mortality and viral loads, as well as more severe pathology caused by excessive production of the inflammatory cytokines. Our studies establish the importance of free ISG15 in IFNα-induced antiviral immunity and in the control of viral infections.
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Affiliation(s)
- Huimin Liu
- College of Life Sciences, Henan Agricultural University, Zhengzhou, Henan, China
| | - Chen Li
- College of Life Sciences, Henan Agricultural University, Zhengzhou, Henan, China
| | - Wenfeng He
- College of Life Sciences, Henan Agricultural University, Zhengzhou, Henan, China
| | - Jing Chen
- College of Life Sciences, Henan Agricultural University, Zhengzhou, Henan, China
| | - Guoqing Yang
- College of Life Sciences, Henan Agricultural University, Zhengzhou, Henan, China
| | - Lu Chen
- College of Veterinary Medicine, Henan Agricultural University, Zhengzhou, Henan, China
| | - Hongtao Chang
- College of Veterinary Medicine, Henan Agricultural University, Zhengzhou, Henan, China
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13
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Interferon-Stimulated Gene 15 Knockout in Mice Impairs IFNα-Mediated Antiviral Activity. Viruses 2022; 14:v14091862. [PMID: 36146669 PMCID: PMC9502845 DOI: 10.3390/v14091862] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/17/2022] [Revised: 08/19/2022] [Accepted: 08/19/2022] [Indexed: 11/25/2022] Open
Abstract
Type I interferon (IFN) plays an important role in the host defense against viral infection by inducing expression of interferon-stimulated genes (ISGs). In a previous study, we found that porcine interferon-stimulated gene 15 (ISG15) exhibited antiviral activity against PRV in vitro. To further investigate the antiviral function of ISG15 in vivo, we utilized ISG15 knockout (ISG15-/-) mice in this study. Here, we demonstrate that ISG15-/- mice were highly susceptible to PRV infection in vivo, as evidenced by a considerably reduced survival rate, enhanced viral replication and severe pathological lesions. However, we observed no significant difference between female and male infected WT and ISG15-/- mice. Moreover, ISG15-/- mice displayed attenuated antiviral protection as a result of considerably reduced expression of IFNβ and relevant ISGs during PRV replication. Furthermore, excessive production of proinflammatory cytokines may be closely related to encephalitis and pneumonia. In further studies, we found that the enhanced sensitivity to PRV infection in ISG15-/- mice might be caused by reduced phosphorylation of STAT1 and STAT2, thereby inhibiting type I IFN-mediated antiviral activity. Based on these findings, we conclude that ISG15 is essential for host type I IFN-mediated antiviral response.
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14
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Wang Z, Li T, Gong Z, Xie J. Role of ISG15 post-translational modification in immunity against Mycobacterium tuberculosis infection. Cell Signal 2022; 94:110329. [PMID: 35390466 DOI: 10.1016/j.cellsig.2022.110329] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/16/2022] [Revised: 03/30/2022] [Accepted: 03/31/2022] [Indexed: 11/30/2022]
Abstract
ISG15 encoded by a type I interferon (IFN) inducible gene mediates an important cellular process called ISGylation. ISGylation emerges as a powerful host tactic against intracellular pathogens like Mycobacterium tuberculosis (Mtb). However, the exact role of ISGylation in immunity remains elusive. To shed light on how ISGylation, which is both interesting and complex, participates in immunity against Mtb, this manuscript summarized the current knowledge about the structural characteristics and targets of ISG15 and how ISGylation cross-talks with other host post-translational modifications to exert its effect.
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Affiliation(s)
- Zilu Wang
- Institute of Modern Biopharmaceuticals, State Key Laboratory Breeding Base of Eco-Environment and Bio-Resource of the Three Gorges Area, Key Laboratory of Eco-environments in Three Gorges Reservoir Region, Ministry of Education, School of Life Sciences, Southwest University, Chongqing 400715, China
| | - Tongxin Li
- Chongqing Public Health Medical Center, Southwest University Public Health Hospital, central laboratory Chongqing, 400030, China
| | - Zhen Gong
- Institute of Modern Biopharmaceuticals, State Key Laboratory Breeding Base of Eco-Environment and Bio-Resource of the Three Gorges Area, Key Laboratory of Eco-environments in Three Gorges Reservoir Region, Ministry of Education, School of Life Sciences, Southwest University, Chongqing 400715, China
| | - Jianping Xie
- Institute of Modern Biopharmaceuticals, State Key Laboratory Breeding Base of Eco-Environment and Bio-Resource of the Three Gorges Area, Key Laboratory of Eco-environments in Three Gorges Reservoir Region, Ministry of Education, School of Life Sciences, Southwest University, Chongqing 400715, China.
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15
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Jurczyszak D, Manganaro L, Buta S, Gruber C, Martin-Fernandez M, Taft J, Patel RS, Cipolla M, Alshammary H, Mulder LCF, Sachidanandam R, Bogunovic D, Simon V. ISG15 deficiency restricts HIV-1 infection. PLoS Pathog 2022; 18:e1010405. [PMID: 35333911 PMCID: PMC8986114 DOI: 10.1371/journal.ppat.1010405] [Citation(s) in RCA: 22] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2021] [Revised: 04/06/2022] [Accepted: 02/28/2022] [Indexed: 01/01/2023] Open
Abstract
Type I interferons (IFN-Is) are a group of potent inflammatory and antiviral cytokines. They induce IFN stimulated genes (ISGs), which act as proinflammatory mediators, antiviral effectors, and negative regulators of the IFN-I signaling cascade itself. One such regulator is interferon stimulated gene 15 (ISG15). Humans with complete ISG15 deficiency express persistently elevated levels of ISGs, and consequently, exhibit broad spectrum resistance to viral infection. Here, we demonstrate that IFN-I primed fibroblasts derived from ISG15-deficient individuals are more resistant to infection with single-cycle HIV-1 compared to healthy control fibroblasts. Complementation with both wild-type (WT) ISG15 and ISG15ΔGG (incapable of ISGylation while retaining negative regulation activity) was sufficient to reverse this phenotype, restoring susceptibility to infection to levels comparable to WT cells. Furthermore, CRISPR-edited ISG15ko primary CD4+ T cells were less susceptible to HIV-1 infection compared to cells treated with non-targeting controls. Transcriptome analysis of these CRISPR-edited ISG15ko primary CD4+ T cells recapitulated the ISG signatures of ISG15 deficient patients. Taken together, we document that the increased broad-spectrum viral resistance in ISG15-deficiency also extends to HIV-1 and is driven by a combination of T-cell-specific ISGs, with both known and unknown functions, predicted to target HIV-1 replication at multiple steps. Type I interferons (IFN-Is) are a group of potent inflammatory and antiviral agents. They induce IFN stimulated genes (ISGs), which perform downstream functions to resolve viral infection, mediate the inflammatory response, as well as negatively regulate the IFN-I signaling cascade to prevent hyperinflammation. One such negative regulator is interferon stimulated gene 15 (ISG15). Humans that lack ISG15 have chronic, low levels of antiviral ISGs, and ensuing broad-spectrum resistance to viral infection. We demonstrate that IFN-I priming of ISG15-deficient cells leads to superior resistance to human immunodeficiency virus 1 (HIV-1) infection compared to IFN-I primed healthy control cells. This is true for fibroblast cell lines, as well as primary CD4+ T cells, the main target of HIV-1. Analysis of the gene expression profiles show that ISG15-knockout CD4+ T cells express similar inflammatory markers as ISG15-deficient patients. Overall, we show that the broad-spectrum viral resistance in ISG15-deficiency extends to HIV-1.
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Affiliation(s)
- Denise Jurczyszak
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Center for Inborn Errors of Immunity, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Department of Pediatrics, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
| | - Lara Manganaro
- INGM-Istituto Nazionale di Genetica Molecolare, Virology, Milan, Italy
- Department of Pharmacological and Biomolecular Sciences (DiSFeB), University of MIlan, Milan, Italy
| | - Sofija Buta
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Center for Inborn Errors of Immunity, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Department of Pediatrics, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
| | - Conor Gruber
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Center for Inborn Errors of Immunity, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Department of Pediatrics, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
| | - Marta Martin-Fernandez
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Center for Inborn Errors of Immunity, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Department of Pediatrics, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
| | - Justin Taft
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Center for Inborn Errors of Immunity, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Department of Pediatrics, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
| | - Roosheel S. Patel
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Center for Inborn Errors of Immunity, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Department of Pediatrics, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
| | - Melissa Cipolla
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Center for Inborn Errors of Immunity, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Department of Pediatrics, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
| | - Hala Alshammary
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
| | - Lubbertus C. F. Mulder
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Global Health and Emerging Pathogens Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
| | - Ravi Sachidanandam
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
| | - Dusan Bogunovic
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Center for Inborn Errors of Immunity, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Precision Immunology Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Department of Pediatrics, Icahn School of Medicine at Mount Sinai, New York city, New York, United States of America
- Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- * E-mail: (DB); (VS)
| | - Viviana Simon
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Global Health and Emerging Pathogens Institute, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- Department of Pathology, Molecular and Cell-Based Medicine, Icahn School of Medicine at Mount Sinai, New York City, New York, United States of America
- * E-mail: (DB); (VS)
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16
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Mirzalieva O, Juncker M, Schwartzenburg J, Desai S. ISG15 and ISGylation in Human Diseases. Cells 2022; 11:cells11030538. [PMID: 35159348 PMCID: PMC8834048 DOI: 10.3390/cells11030538] [Citation(s) in RCA: 52] [Impact Index Per Article: 17.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/01/2021] [Revised: 01/18/2022] [Accepted: 01/25/2022] [Indexed: 12/04/2022] Open
Abstract
Type I Interferons (IFNs) induce the expression of >500 genes, which are collectively called ISGs (IFN-stimulated genes). One of the earliest ISGs induced by IFNs is ISG15 (Interferon-Stimulated Gene 15). Free ISG15 protein synthesized from the ISG15 gene is post-translationally conjugated to cellular proteins and is also secreted by cells into the extracellular milieu. ISG15 comprises two ubiquitin-like domains (UBL1 and UBL2), each of which bears a striking similarity to ubiquitin, accounting for its earlier name ubiquitin cross-reactive protein (UCRP). Like ubiquitin, ISG15 harbors a characteristic β-grasp fold in both UBL domains. UBL2 domain has a conserved C-terminal Gly-Gly motif through which cellular proteins are appended via an enzymatic cascade similar to ubiquitylation called ISGylation. ISG15 protein is minimally expressed under physiological conditions. However, its IFN-dependent expression is aberrantly elevated or compromised in various human diseases, including multiple types of cancer, neurodegenerative disorders (Ataxia Telangiectasia and Amyotrophic Lateral Sclerosis), inflammatory diseases (Mendelian Susceptibility to Mycobacterial Disease (MSMD), bacteriopathy and viropathy), and in the lumbar spinal cords of veterans exposed to Traumatic Brain Injury (TBI). ISG15 and ISGylation have both inhibitory and/or stimulatory roles in the etiology and pathogenesis of human diseases. Thus, ISG15 is considered a “double-edged sword” for human diseases in which its expression is elevated. Because of the roles of ISG15 and ISGylation in cancer cell proliferation, migration, and metastasis, conferring anti-cancer drug sensitivity to tumor cells, and its elevated expression in cancer, neurodegenerative disorders, and veterans exposed to TBI, both ISG15 and ISGylation are now considered diagnostic/prognostic biomarkers and therapeutic targets for these ailments. In the current review, we shall cover the exciting journey of ISG15, spanning three decades from the bench to the bedside.
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Affiliation(s)
| | | | | | - Shyamal Desai
- Correspondence: ; Tel.: +1-504-568-4388; Fax: +1-504-568-2093
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17
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Gao N, Me R, Dai C, Yu FSX. ISG15 Acts as a Mediator of Innate Immune Response to Pseudomonas aeruginosa Infection in C57BL/6J Mouse Corneas. Invest Ophthalmol Vis Sci 2020; 61:26. [PMID: 32416603 PMCID: PMC7405721 DOI: 10.1167/iovs.61.5.26] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/27/2019] [Accepted: 03/30/2020] [Indexed: 01/04/2023] Open
Abstract
Purpose IFN-stimulated gene (ISG) 15 is a type 1 IFN-induced protein and known to modify target proteins in a manner similar to ubiquitylation (protein conjugation by ISG15 is termed ISGylation). We sought to determine the role of ISG15 and its underlying mechanisms in corneal innate immune defense against Pseudomonas aeruginosa keratitis. Methods ISG15 expression in cultured human corneal epithelial cells (HCECs) and mouse corneas was determined by PCR and Western blot analysis. Gene knockout mice were used to define the role of ISG15 signaling in controlling the severity of P. aeruginosa keratitis, which was assessed with photographing, clinical scoring, bacterial counting, myeloperoxidase assay, and quantitative PCR determination of cytokine expression. Integrin LFA-1 inhibitor was used to assess its involvement of ISG15 signaling in P. aeruginosa-infected corneas. Results Heat-killed P. aeruginosa induced ISG15 expression in cultured HCECs and accumulation in the conditioned media. Isg15 deficiency accelerated keratitis progress, suppressed IFNγ and CXCL10, and promoted IL-1β while exhibiting no effects on IFNα expression. Moreover, exogenous ISG15 protected the corneas of wild-type mice from P. aeruginosa infection while markedly reducing the severity of P. aeruginosa keratitis in type 1 IFN-receptor knockout mice. Exogenous ISG15 increased bacteriostatic activity of B6 mouse corneal homogenates, and inhibition of LFA-1 exacerbated the severity of and abolished protective effects of ISG15 on P. aeruginosa keratitis. Conclusions Type 1 INF-induced ISG15 regulates the innate immune response and greatly reduces the susceptibility of B6 mouse corneas to P. aeruginosa infection in an LFA-1-dependent manner.
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18
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Kespohl M, Bredow C, Klingel K, Voß M, Paeschke A, Zickler M, Poller W, Kaya Z, Eckstein J, Fechner H, Spranger J, Fähling M, Wirth EK, Radoshevich L, Thery F, Impens F, Berndt N, Knobeloch KP, Beling A. Protein modification with ISG15 blocks coxsackievirus pathology by antiviral and metabolic reprogramming. SCIENCE ADVANCES 2020; 6:eaay1109. [PMID: 32195343 PMCID: PMC7065878 DOI: 10.1126/sciadv.aay1109] [Citation(s) in RCA: 31] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/21/2019] [Accepted: 12/13/2019] [Indexed: 05/10/2023]
Abstract
Protein modification with ISG15 (ISGylation) represents a major type I IFN-induced antimicrobial system. Common mechanisms of action and species-specific aspects of ISGylation, however, are still ill defined and controversial. We used a multiphasic coxsackievirus B3 (CV) infection model with a first wave resulting in hepatic injury of the liver, followed by a second wave culminating in cardiac damage. This study shows that ISGylation sets nonhematopoietic cells into a resistant state, being indispensable for CV control, which is accomplished by synergistic activity of ISG15 on antiviral IFIT1/3 proteins. Concurrent with altered energy demands, ISG15 also adapts liver metabolism during infection. Shotgun proteomics, in combination with metabolic network modeling, revealed that ISG15 increases the oxidative capacity and promotes gluconeogenesis in liver cells. Cells lacking the activity of the ISG15-specific protease USP18 exhibit increased resistance to clinically relevant CV strains, therefore suggesting that stabilizing ISGylation by inhibiting USP18 could be exploited for CV-associated human pathologies.
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Affiliation(s)
- Meike Kespohl
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Biochemistry, Berlin, Germany
- Deutsches Zentrum für Herz-Kreislauf-Forschung (DZHK), partner site Berlin, Germany
| | - Clara Bredow
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Biochemistry, Berlin, Germany
| | - Karin Klingel
- University of Tuebingen, Cardiopathology, Institute for Pathology and Neuropathology, Tuebingen, Germany
| | - Martin Voß
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Biochemistry, Berlin, Germany
| | - Anna Paeschke
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Biochemistry, Berlin, Germany
| | - Martin Zickler
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Biochemistry, Berlin, Germany
| | - Wolfgang Poller
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Clinic for Cardiology, Campus Benjamin Franklin, Berlin, Germany
| | - Ziya Kaya
- Universitätsklinikum Heidelberg, Medizinische Klinik für Innere Medizin III: Kardiologie, Angiologie und Pneumologie, Heidelberg, Germany
- Deutsches Zentrum für Herz-Kreislauf-Forschung (DZHK), partner site Heidelberg, Germany
| | - Johannes Eckstein
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Biochemistry, Berlin, Germany
| | - Henry Fechner
- Department of Applied Biochemistry, Institute of Biotechnology, Technische Universität Berlin, Berlin, Germany
| | - Joachim Spranger
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Department of Endocrinology, Diabetes and Nutrition, Berlin, Germany
| | - Michael Fähling
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Vegetative Physiology, Berlin, Germany
| | - Eva Katrin Wirth
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Department of Endocrinology, Diabetes and Nutrition, Berlin, Germany
| | - Lilliana Radoshevich
- Department of Microbiology and Immunology, University of Iowa, Iowa City, Iowa, USA
| | - Fabien Thery
- Department of Biomolecular Medicine, Ghent University, Ghent, Belgium
- VIB Center for Medical Biotechnology, Ghent, Belgium
| | - Francis Impens
- Department of Biomolecular Medicine, Ghent University, Ghent, Belgium
- VIB Center for Medical Biotechnology, Ghent, Belgium
- VIB Proteomics Core, Ghent, Belgium
| | - Nikolaus Berndt
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Biochemistry, Berlin, Germany
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute for Computational and Imaging Science in Cardiovascular Medicine, Berlin, Germany
| | | | - Antje Beling
- Charité—Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Biochemistry, Berlin, Germany
- Deutsches Zentrum für Herz-Kreislauf-Forschung (DZHK), partner site Berlin, Germany
- Corresponding author.
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19
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Clasman JR, Everett RK, Srinivasan K, Mesecar AD. Decoupling deISGylating and deubiquitinating activities of the MERS virus papain-like protease. Antiviral Res 2020; 174:104661. [PMID: 31765674 PMCID: PMC7114298 DOI: 10.1016/j.antiviral.2019.104661] [Citation(s) in RCA: 38] [Impact Index Per Article: 7.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2019] [Revised: 11/12/2019] [Accepted: 11/18/2019] [Indexed: 11/17/2022]
Abstract
Coronavirus papain-like proteases (PLPs or PLpro), such as the one encoded in the genome of the infectious Middle East Respiratory Syndrome (MERS) virus, have multiple enzymatic activities that promote viral infection. PLpro acts as a protease and processes the large coronavirus polyprotein for virus replication. PLpro also functions as both a deubiquitinating (DUB) and deISGylating (deISG) enzyme and removes ubiquitin (Ub) and interferon-stimulated gene 15 (ISG15) from cellular proteins. Both DUB and deISG activities are implicated in suppressing innate immune responses; however, the precise role of each activity in this process is still unclear due in part to the difficulties in separating each activity. In this study, we determine the first structure of MERS PLpro in complex with the full-length human ISG15 to a resolution of 2.3 Å. This structure and available structures of MERS PLpro-Ub complexes were used as molecular guides to design PLpro mutants that lack either or both DUB/deISG activities. We tested 13 different PLpro mutants for protease, DUB, and deISG activitites using fluorescence-based assays. Results show that we can selectively modulate DUB activity at amino acid positions 1649 and 1653 while mutation of Val1691 or His1652 of PLpro to a positive charged residue completely impairs both DUB/deISG activities. These mutant enzymes will provide new functional tools for delineating the importance of DUB versus deISG activity in virus-infected cells and may serve as potential candidates for attenuating the MERS virus in vivo for modified vaccine design efforts.
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Affiliation(s)
- Jozlyn R Clasman
- Department of Biological Sciences, Purdue University, West Lafayette, IN, USA
| | - Renata K Everett
- Department of Biological Sciences, Purdue University, West Lafayette, IN, USA
| | - Karthik Srinivasan
- Department of Biological Sciences, Purdue University, West Lafayette, IN, USA
| | - Andrew D Mesecar
- Department of Biological Sciences, Purdue University, West Lafayette, IN, USA; Department of Biochemistry, Purdue University, West Lafayette, IN, USA; Center for Cancer Research, Purdue University, West Lafayette, IN, USA.
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20
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Zhang Y, Thery F, Wu NC, Luhmann EK, Dussurget O, Foecke M, Bredow C, Jiménez-Fernández D, Leandro K, Beling A, Knobeloch KP, Impens F, Cossart P, Radoshevich L. The in vivo ISGylome links ISG15 to metabolic pathways and autophagy upon Listeria monocytogenes infection. Nat Commun 2019; 10:5383. [PMID: 31772204 PMCID: PMC6879477 DOI: 10.1038/s41467-019-13393-x] [Citation(s) in RCA: 62] [Impact Index Per Article: 10.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2019] [Accepted: 11/07/2019] [Indexed: 12/28/2022] Open
Abstract
ISG15 is an interferon-stimulated, ubiquitin-like protein, with anti-viral and anti-bacterial activity. Here, we map the endogenous in vivo ISGylome in the liver following Listeria monocytogenes infection by combining murine models of reduced or enhanced ISGylation with quantitative proteomics. Our method identifies 930 ISG15 sites in 434 proteins and also detects changes in the host ubiquitylome. The ISGylated targets are enriched in proteins which alter cellular metabolic processes, including upstream modulators of the catabolic and antibacterial pathway of autophagy. Computational analysis of substrate structures reveals that a number of ISG15 modifications occur at catalytic sites or dimerization interfaces of enzymes. Finally, we demonstrate that animals and cells with enhanced ISGylation have increased basal and infection-induced autophagy through the modification of mTOR, WIPI2, AMBRA1, and RAB7. Taken together, these findings ascribe a role of ISGylation to temporally reprogram organismal metabolism following infection through direct modification of a subset of enzymes in the liver.
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Affiliation(s)
- Yifeng Zhang
- Department of Microbiology and Immunology, University of Iowa Carver College of Medicine, Iowa City, IA, 52242, USA
| | - Fabien Thery
- Center for Medical Biotechnology, VIB, 9000, Gent, Belgium
- Department for Biomolecular Medicine, Gent University, 9000, Gent, Belgium
| | - Nicholas C Wu
- Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA, 92037, USA
| | - Emma K Luhmann
- Department of Microbiology and Immunology, University of Iowa Carver College of Medicine, Iowa City, IA, 52242, USA
| | - Olivier Dussurget
- Institut Pasteur, Unité des Interactions Bactéries-Cellules, Département de Biologie Cellulaire et Infection, 75015, Paris, France
- Inserm, U604, 75015, Paris, France
- National Institute for Agronomic Research (INRA), Unité sous-contrat 2020, 75015, Paris, France
| | - Mariko Foecke
- Institut Pasteur, Unité des Interactions Bactéries-Cellules, Département de Biologie Cellulaire et Infection, 75015, Paris, France
- Inserm, U604, 75015, Paris, France
- National Institute for Agronomic Research (INRA), Unité sous-contrat 2020, 75015, Paris, France
| | - Clara Bredow
- Charité-Universitäts medizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Biochemistry, Berlin, Germany
| | | | - Kevin Leandro
- Center for Medical Biotechnology, VIB, 9000, Gent, Belgium
- Department for Biomolecular Medicine, Gent University, 9000, Gent, Belgium
| | - Antje Beling
- Charité-Universitäts medizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health (BIH), Institute of Biochemistry, Berlin, Germany
- Deutsches Zentrum für Herz-Kreislauf-Forschung (DZHK), Partner Site Berlin, Berlin, Germany
| | - Klaus-Peter Knobeloch
- Institute of Neuropathology, Medical Faculty, University of Freiburg, Freiburg, Germany
| | - Francis Impens
- Center for Medical Biotechnology, VIB, 9000, Gent, Belgium.
- Department for Biomolecular Medicine, Gent University, 9000, Gent, Belgium.
- VIB Proteomics Core, VIB, 9000, Gent, Belgium.
| | - Pascale Cossart
- Institut Pasteur, Unité des Interactions Bactéries-Cellules, Département de Biologie Cellulaire et Infection, 75015, Paris, France.
- Inserm, U604, 75015, Paris, France.
- National Institute for Agronomic Research (INRA), Unité sous-contrat 2020, 75015, Paris, France.
| | - Lilliana Radoshevich
- Department of Microbiology and Immunology, University of Iowa Carver College of Medicine, Iowa City, IA, 52242, USA.
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21
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Li J, Johnson JA, Su H. Ubiquitin and Ubiquitin-like proteins in cardiac disease and protection. Curr Drug Targets 2019; 19:989-1002. [PMID: 26648080 DOI: 10.2174/1389450117666151209114608] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2015] [Accepted: 11/01/2015] [Indexed: 01/10/2023]
Abstract
Post-translational modification represents an important mechanism to regulate protein function in cardiac cells. Ubiquitin (Ub) and ubiquitin-like proteins (UBLs) are a family of protein modifiers that share a certain extent of sequence and structure similarity. Conjugation of Ub or UBLs to target proteins is dynamically regulated by a set of UBL-specific enzymes and modulates the physical and physiological properties of protein substrates. Ub and UBLs control a strikingly wide spectrum of cellular processes and not surprisingly are involved in the development of multiple human diseases including cardiac diseases. Further identification of novel UBL targets will expand our understanding of the functional diversity of UBL pathways in physiology and pathology. Here we review recent findings on the mechanisms, proteome and functions of a subset of UBLs and highlight their potential impacts on the development and progression of various forms of cardiac diseases.
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Affiliation(s)
- Jie Li
- Vascular Biology Center, Medical College of Georgia, Augusta University, Augusta, GA, United States
| | - John A Johnson
- Department of Pharmacology and Toxicology, Medical College of Georgia, Augusta University, Augusta, GA, United States
| | - Huabo Su
- Vascular Biology Center, Medical College of Georgia, Augusta University, Augusta, GA, United States.,Department of Pharmacology and Toxicology, Medical College of Georgia, Augusta University, Augusta, GA, United States
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22
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Loss of TRIM29 suppresses cancer stem cell-like characteristics of PDACs via accelerating ISG15 degradation. Oncogene 2019; 39:546-559. [DOI: 10.1038/s41388-019-0992-2] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/06/2018] [Revised: 06/13/2019] [Accepted: 06/15/2019] [Indexed: 12/30/2022]
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23
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Abstract
The host response to viral infection includes the induction of type I interferons and the subsequent upregulation of hundreds of interferon-stimulated genes. Ubiquitin-like protein ISG15 is an interferon-induced protein that has been implicated as a central player in the host antiviral response. Over the past 15 years, efforts to understand how ISG15 protects the host during infection have revealed that its actions are diverse and pathogen-dependent. In this Review, we describe new insights into how ISG15 directly inhibits viral replication and discuss the recent finding that ISG15 modulates the host damage and repair response, immune response and other host signalling pathways. We also explore the viral immune-evasion strategies that counteract the actions of ISG15. These findings are integrated with a discussion of the recent identification of ISG15-deficient individuals and a cellular receptor for ISG15 that provides new insights into how ISG15 shapes the host response to viral infection. Ubiquitin-like protein ISG15 is an interferon-induced protein that has been implicated as a central player in the host antiviral response. In this Review, Perng and Lenschow provide new insights into how ISG15 restricts and shapes the host response to viral infection and the viral immune-evasion strategies that counteract ISG15.
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Affiliation(s)
- Yi-Chieh Perng
- Department of Internal Medicine, Washington University School of Medicine, St Louis, MO, USA
| | - Deborah J Lenschow
- Department of Internal Medicine, Washington University School of Medicine, St Louis, MO, USA. .,Department of Pathology and Immunology, Washington University School of Medicine, St Louis, MO, USA.
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24
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BAG3 deletion suppresses stem cell-like features of pancreatic ductal adenocarcinoma via translational suppression of ISG15. BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 2019; 1866:819-827. [DOI: 10.1016/j.bbamcr.2019.02.008] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/19/2018] [Revised: 02/12/2019] [Accepted: 02/13/2019] [Indexed: 11/21/2022]
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25
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Albert M, Bécares M, Falqui M, Fernández-Lozano C, Guerra S. ISG15, a Small Molecule with Huge Implications: Regulation of Mitochondrial Homeostasis. Viruses 2018; 10:v10110629. [PMID: 30428561 PMCID: PMC6265978 DOI: 10.3390/v10110629] [Citation(s) in RCA: 37] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2018] [Revised: 11/08/2018] [Accepted: 11/09/2018] [Indexed: 12/12/2022] Open
Abstract
Viruses are responsible for the majority of infectious diseases, from the common cold to HIV/AIDS or hemorrhagic fevers, the latter with devastating effects on the human population. Accordingly, the development of efficient antiviral therapies is a major goal and a challenge for the scientific community, as we are still far from understanding the molecular mechanisms that operate after virus infection. Interferon-stimulated gene 15 (ISG15) plays an important antiviral role during viral infection. ISG15 catalyzes a ubiquitin-like post-translational modification termed ISGylation, involving the conjugation of ISG15 molecules to de novo synthesized viral or cellular proteins, which regulates their stability and function. Numerous biomedically relevant viruses are targets of ISG15, as well as proteins involved in antiviral immunity. Beyond their role as cellular powerhouses, mitochondria are multifunctional organelles that act as signaling hubs in antiviral responses. In this review, we give an overview of the biological consequences of ISGylation for virus infection and host defense. We also compare several published proteomic studies to identify and classify potential mitochondrial ISGylation targets. Finally, based on our recent observations, we discuss the essential functions of mitochondria in the antiviral response and examine the role of ISG15 in the regulation of mitochondrial processes, specifically OXPHOS and mitophagy.
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Affiliation(s)
- Manuel Albert
- Department of Preventive Medicine, Public Health and Microbiology, Universidad Autónoma, E-28029 Madrid, Spain.
| | - Martina Bécares
- Department of Preventive Medicine, Public Health and Microbiology, Universidad Autónoma, E-28029 Madrid, Spain.
| | - Michela Falqui
- Department of Preventive Medicine, Public Health and Microbiology, Universidad Autónoma, E-28029 Madrid, Spain.
| | - Carlos Fernández-Lozano
- Department of Preventive Medicine, Public Health and Microbiology, Universidad Autónoma, E-28029 Madrid, Spain.
| | - Susana Guerra
- Department of Preventive Medicine, Public Health and Microbiology, Universidad Autónoma, E-28029 Madrid, Spain.
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Abstract
Alphaviruses, members of the positive-sense, single-stranded RNA virus family Togaviridae, represent a re-emerging public health concern worldwide as mosquito vectors expand into new geographic ranges. Members of the alphavirus genus tend to induce clinical disease characterized by rash, arthralgia, and arthritis (chikungunya virus, Ross River virus, and Semliki Forest virus) or encephalomyelitis (eastern equine encephalitis virus, western equine encephalitis virus, and Venezuelan equine encephalitis virus), though some patients who recover from the initial acute illness may develop long-term sequelae, regardless of the specific infecting virus. Studies examining the natural disease course in humans and experimental infection in cell culture and animal models reveal that host genetics play a major role in influencing susceptibility to infection and severity of clinical disease. Genome-wide genetic screens, including loss of function screens, microarrays, RNA-sequencing, and candidate gene studies, have further elucidated the role host genetics play in the response to virus infection, with the immune response being found in particular to majorly influence the outcome. This review describes the current knowledge of the mechanisms by which host genetic factors influence alphavirus pathogenesis and discusses emerging technologies that are poised to increase our understanding of the complex interplay between viral and host genetics on disease susceptibility and clinical outcome.
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Evolution-Guided Structural and Functional Analyses of the HERC Family Reveal an Ancient Marine Origin and Determinants of Antiviral Activity. J Virol 2018; 92:JVI.00528-18. [PMID: 29669830 PMCID: PMC6002735 DOI: 10.1128/jvi.00528-18] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/28/2018] [Accepted: 04/10/2018] [Indexed: 01/24/2023] Open
Abstract
In humans, homologous to the E6-AP carboxyl terminus (HECT) and regulator of chromosome condensation 1 (RCC1)-like domain-containing protein 5 (HERC5) is an interferon-induced protein that inhibits replication of evolutionarily diverse viruses, including human immunodeficiency virus type 1 (HIV-1). To better understand the origin, evolution, and function of HERC5, we performed phylogenetic, structural, and functional analyses of the entire human small-HERC family, which includes HERC3, HERC4, HERC5, and HERC6. We demonstrated that the HERC family emerged >595 million years ago and has undergone gene duplication and gene loss events throughout its evolution. The structural topology of the RCC1-like domain and HECT domains from all HERC paralogs is highly conserved among evolutionarily diverse vertebrates despite low sequence homology. Functional analyses showed that the human small HERCs exhibit different degrees of antiviral activity toward HIV-1 and that HERC5 provides the strongest inhibition. Notably, coelacanth HERC5 inhibited simian immunodeficiency virus (SIV), but not HIV-1, particle production, suggesting that the antiviral activity of HERC5 emerged over 413 million years ago and exhibits species- and virus-specific restriction. In addition, we showed that both HERC5 and HERC6 are evolving under strong positive selection, particularly blade 1 of the RCC1-like domain, which we showed is a key determinant of antiviral activity. These studies provide insight into the origin, evolution, and biological importance of the human restriction factor HERC5 and the other HERC family members. IMPORTANCE Intrinsic immunity plays an important role as the first line of defense against viruses. Studying the origins, evolution, and functions of proteins responsible for effecting this defense will provide key information about virus-host relationships that can be exploited for future drug development. We showed that HERC5 is one such antiviral protein that belongs to an evolutionarily conserved family of HERCs with an ancient marine origin. Not all vertebrates possess all HERC members, suggesting that different HERCs emerged at different times during evolution to provide the host with a survival advantage. Consistent with this, two of the more recently emerged HERC members, HERC5 and HERC6, displayed strong signatures of having been involved in an ancient evolutionary battle with viruses. Our findings provide new insights into the evolutionary origin and function of the HERC family in vertebrate evolution, identifying HERC5 and possibly HERC6 as important effectors of intrinsic immunity in vertebrates.
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Wang M, Huang Y, He M, Peng WJ, Tian DY. Effects of hepatitis E virus infection on interferon production via ISG15. World J Gastroenterol 2018; 24:2173-2180. [PMID: 29853735 PMCID: PMC5974579 DOI: 10.3748/wjg.v24.i20.2173] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/29/2018] [Revised: 04/19/2018] [Accepted: 04/23/2018] [Indexed: 02/06/2023] Open
Abstract
AIM To assess the effects of hepatitis E virus (HEV) on the production of type I interferons (IFNs) and determine the underlying mechanisms.
METHODS We measured the production of interferon (IFN)-alpha and -beta (-α/β) in genotype 3 HEV-infected C3A cells at different time points (0, 8, 12, 24, 48, 72 and 120 h) by enzyme-linked immunosorbent assay (ELISA). The expression levels of IFN-stimulated gene (ISG)15 in HEV-infected C3A cells at different time points were tested by western blotting. The plasmid-expressing open reading frame 3 (ORF3) or control plasmids (green fluorescent protein-expressing) were transfected into C3A cells, and the levels of IFN-α/β and ISG15 were evaluated, respectively. Furthermore, the plasmid-expressing ISG15 or small interfering RNA-inhibiting ISG15 was transfected into infected C3A cells. Then, the production of IFN-α/β was also measured by ELISA.
RESULTS We showed that genotype 3 HEV could enhance the production of IFN-α/β and induce elevation of ISG15 in C3A cells. HEV ORF3 protein could enhance the production of IFN-α/β and the expression of ISG15. Additionally, ISG15 silencing enhanced the production of IFN-α/β. Overexpression of ISG15 resulted in the reduction of IFN-α/β.
CONCLUSION HEV may promote production of IFN-α/β and expression of ISG15 via ORF3 in the early stages, and increased ISG15 subsequently inhibited the production of IFN-α/β.
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Affiliation(s)
- Min Wang
- Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, Hubei Province, China
| | - Ying Huang
- The First Affiliated Hospital of Sun Yat-sen University, Guangzhou 510700, Guangdong Province, China
| | - Man He
- Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, Hubei Province, China
| | - Wen-Ju Peng
- Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, Hubei Province, China
| | - De-Ying Tian
- Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, Hubei Province, China
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Zhao P, Jiang T, Zhong Z, Zhao L, Yang S, Xia X. Inhibition of rabies virus replication by interferon-stimulated gene 15 and its activating enzyme UBA7. INFECTION GENETICS AND EVOLUTION 2017; 56:44-53. [PMID: 29056542 DOI: 10.1016/j.meegid.2017.10.016] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/13/2017] [Accepted: 10/19/2017] [Indexed: 01/27/2023]
Abstract
It was reported that ISG15 and its activating enzyme UBA7 have antiviral functions. However, there is no study to demonstrate whether ISG15 and UBA7 have anti-rabies virus function. In the current study, In vivo and in vitro anti-rabies virus function of ISG15 and UBA7 were investigated using RNAi technology. The results showed that shRNA knock-down of expression of ISG15 and UBA7 increased the titers of RABV in neuroblastoma cell line NA and microglial cell line BV-2 cells and shRNA knockdown of ISG15 conjugation alleviates the IFN-induced inhibition of RABV gene expression in vitro. Lentiviral vector mediated-shRNA knock-down of expression of ISG15 and UBA7 increased the titers of RABV in mouse brains and decreased the survivorship of mice. The study showed that ISG15 and UBA7 inhibit RABV replication in vitro and in vivo. To our knowledge, we for the first time documented the anti-RABV function of ISG15 and UBA7, which may provide a means of understanding the pathogenesis of rabies and improving therapeutic methods.
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Affiliation(s)
- Pingsen Zhao
- Clinical Core Laboratory, Meizhou People's Hospital (Huangtang Hospital), Meizhou Hospital Affiliated to Sun Yat-sen University, Meizhou 514031, PR China; Center for Precision Medicine, Meizhou People's Hospital (Huangtang Hospital), Meizhou Hospital Affiliated to Sun Yat-sen University, Meizhou 514031, PR China.
| | - Tianqi Jiang
- College of Veterinary Medicine, Northeast Agricultural University, Harbin 150030, China
| | - Zhixiong Zhong
- Center for Precision Medicine, Meizhou People's Hospital (Huangtang Hospital), Meizhou Hospital Affiliated to Sun Yat-sen University, Meizhou 514031, PR China
| | - Lili Zhao
- Institute of Military Veterinary, Academy of Military Medical Sciences, Changchun 130122, China
| | - Songtao Yang
- Institute of Military Veterinary, Academy of Military Medical Sciences, Changchun 130122, China
| | - Xianzhu Xia
- Institute of Military Veterinary, Academy of Military Medical Sciences, Changchun 130122, China
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30
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UBE2L6/UBCH8 and ISG15 attenuate autophagy in esophageal cancer cells. Oncotarget 2017; 8:23479-23491. [PMID: 28186990 PMCID: PMC5410320 DOI: 10.18632/oncotarget.15182] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2016] [Accepted: 01/16/2017] [Indexed: 12/22/2022] Open
Abstract
Esophageal cancer remains a poor prognosis cancer due to advanced stage of presentation and drug resistant disease. To understand the molecular mechanisms influencing response to chemotherapy, we examined genes that are differentially expressed between drug sensitive, apoptosis competent esophageal cancer cells (OE21, OE33, FLO-1) and those which are more resistant and do not exhibit apoptosis (KYSE450 and OE19). Members of the ISG15 (ubiquitin-like) protein modification pathway, including UBE2L6 and ISG15, were found to be more highly expressed in the drug sensitive cell lines. In this study, we evaluated the contribution of these proteins to the response of drug sensitive cells. Depletion of UBE2L6 or ISG15 with siRNA did not influence caspase-3 activation or nuclear fragmentation following treatment with 5-fluorouracil (5-FU). We assessed autophagy by analysis of LC3II expression and Cyto-ID staining. Depletion of either ISG15 or UBE2L6 resulted in enhanced endogenous autophagic flux. An increase in autophagic flux was also observed following treatment with cytotoxic drugs (5-FU, rapamycin). In ISG15 depleted cells, this increase in autophagy was associated with improved recovery of drug treated cells. In contrast, UBE2L6 depleted cells, did not show enhanced recovery. UBE2L6 may therefore influence additional targets that limit the pro-survival effect of ISG15 depletion. These data identify UBE2L6 and ISG15 as novel inhibitors of autophagy, with the potential to influence chemosensitivity in esophageal cancer cells.
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31
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Manry J, Nédélec Y, Fava VM, Cobat A, Orlova M, Thuc NV, Thai VH, Laval G, Barreiro LB, Schurr E. Deciphering the genetic control of gene expression following Mycobacterium leprae antigen stimulation. PLoS Genet 2017; 13:e1006952. [PMID: 28793313 PMCID: PMC5565194 DOI: 10.1371/journal.pgen.1006952] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/07/2017] [Revised: 08/21/2017] [Accepted: 08/02/2017] [Indexed: 12/02/2022] Open
Abstract
Leprosy is a human infectious disease caused by Mycobacterium leprae. A strong host genetic contribution to leprosy susceptibility is well established. However, the modulation of the transcriptional response to infection and the mechanism(s) of disease control are poorly understood. To address this gap in knowledge of leprosy pathogenicity, we conducted a genome-wide search for expression quantitative trait loci (eQTL) that are associated with transcript variation before and after stimulation with M. leprae sonicate in whole blood cells. We show that M. leprae antigen stimulation mainly triggered the upregulation of immune related genes and that a substantial proportion of the differential gene expression is genetically controlled. Indeed, using stringent criteria, we identified 318 genes displaying cis-eQTL at an FDR of 0.01, including 66 genes displaying response-eQTL (reQTL), i.e. cis-eQTL that showed significant evidence for interaction with the M. leprae stimulus. Such reQTL correspond to regulatory variations that affect the interaction between human whole blood cells and M. leprae sonicate and, thus, likely between the human host and M. leprae bacilli. We found that reQTL were significantly enriched among binding sites of transcription factors that are activated in response to infection, and that they were enriched among single nucleotide polymorphisms (SNPs) associated with susceptibility to leprosy per se and Type-I Reaction, and seven of them have been targeted by recent positive selection. Our study suggested that natural selection shaped our genomic diversity to face pathogen exposure including M. leprae infection. Each year, 200,000 new leprosy cases are reported worldwide. While there is unambiguous evidence for a role of host genetics in leprosy pathogenesis, the mechanisms by which the human host fights the infection are poorly understood. Here, we highlight the search for naturally occurring genetic variations that modulate gene expression levels following exposure to sonicate of Mycobacterium leprae, the bacterium causing the disease. Because M. leprae is not cultivable and the genuine immune cells involved in the host response during infection are still unknown, we performed a genome-wide search for such genetic variations after stimulation of whole-blood from leprosy patients with M. leprae sonicate. This design allowed to provide a general framework for the genetic control of host responses to M. leprae and outlined the contribution of host genetics to leprosy pathogenesis. Among the M. leprae-dependent genetic regulators of gene expression levels there was an enrichment of variants (i) associated with leprosy, (ii) located in transcription factor binding sites and (iii) targeted by recent positive selection.
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Affiliation(s)
- Jérémy Manry
- Program in Infectious Diseases and Immunity in Global Health, The Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada
- McGill International TB Centre, McGill University, Montreal, Quebec, Canada
- Departments of Medicine and Human Genetics, McGill University, Montreal, Quebec, Canada
- * E-mail: (ES); (JM)
| | - Yohann Nédélec
- Department of Genetics, CHU Sainte-Justine Research Centre, Montreal, Quebec, Canada
- Department of Biochemistry, Faculty of Medicine, University of Montreal, Montreal, Quebec, Canada
| | - Vinicius M. Fava
- Program in Infectious Diseases and Immunity in Global Health, The Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada
- McGill International TB Centre, McGill University, Montreal, Quebec, Canada
- Departments of Medicine and Human Genetics, McGill University, Montreal, Quebec, Canada
| | - Aurélie Cobat
- Laboratory of Human Genetics of Infectious Diseases, Necker Branch, Institut National de la Santé et de la Recherche Médicale U.1163, Paris, France
- Paris Descartes University, Imagine Institute, Paris, France
| | - Marianna Orlova
- Program in Infectious Diseases and Immunity in Global Health, The Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada
- McGill International TB Centre, McGill University, Montreal, Quebec, Canada
- Departments of Medicine and Human Genetics, McGill University, Montreal, Quebec, Canada
| | | | - Vu Hong Thai
- Hospital for Dermato-Venerology, Ho Chi Minh City, Vietnam
| | - Guillaume Laval
- Institut Pasteur, Unit of Human Evolutionary Genetics, Department of Genomes and Genetics, Paris, France
- Centre National de la Recherche Scientifique, URA3012, Paris, France
- Center of Bioinformatics, Biostatistics and Integrative Biology, Institut Pasteur, Paris, France
| | - Luis B. Barreiro
- Department of Genetics, CHU Sainte-Justine Research Centre, Montreal, Quebec, Canada
- Department of Pediatrics, Faculty of Medicine, University of Montreal, Montreal, Quebec, Canada
| | - Erwin Schurr
- Program in Infectious Diseases and Immunity in Global Health, The Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada
- McGill International TB Centre, McGill University, Montreal, Quebec, Canada
- Departments of Medicine and Human Genetics, McGill University, Montreal, Quebec, Canada
- * E-mail: (ES); (JM)
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Przanowski P, Loska S, Cysewski D, Dabrowski M, Kaminska B. ISG'ylation increases stability of numerous proteins including Stat1, which prevents premature termination of immune response in LPS-stimulated microglia. Neurochem Int 2017; 112:227-233. [PMID: 28774718 DOI: 10.1016/j.neuint.2017.07.013] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2016] [Revised: 07/19/2017] [Accepted: 07/29/2017] [Indexed: 01/26/2023]
Abstract
Microglia are myeloid cells in the central nervous system which maintain homeostasis and contribute to repair, but instigate neuroinflammation when are activated by infection, trauma or neurological diseases. Initiation of acute inflammatory responses could be mimicked in vitro by stimulation of microglial cultures with lipopolysaccharide (LPS). We have previously demonstrated Stat-dependent induction of the Uba7 mRNA expression in LPS stimulated microglia. Uba7 is an E1 enzyme crucial for posttranslational protein modifications. ISG'ylation is a process in which ISG15 is covalently attached to lysines of target proteins via the sequential action of three enzymes: the E1-activating enzyme UbE1L (UBA7), the E2-conjugating enzyme UBCH8, and E3 ligase HERC5. Here we use quantitative labeled-free mass spectrometry and gene silencing to determine the role of ISG'ylation in LPS-stimulated microglia. We found the increased mRNA levels of Isg15, Uba7, Ube2l6, Herc6 and profound ISG'ylation in inflammatory microglia. Silencing of Uba7 in BV2 microglial cells results in a profound decrease in the level of hundreds proteins as measured by mass spectrometry. There is statistically significant intersection of Uba7-dependent proteins in LPS-stimulated microglia and three datasets of ISG'ylated proteins reported in earlier studies. Stat1, a main activator of Uba7 expression, was modified by ISG15 after LPS stimulation. The level of both total and phospho-Stat1 is decreased after Uba7 knockdown leading to premature termination of immune responses as evidenced by the reduction of iNos and Ccl5 expression. Our results suggest that increased ISG'ylation in LPS-stimulated microglia supports stability of proteins, including Stat1, which prevents termination of immune responses during inflammation.
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Affiliation(s)
- Piotr Przanowski
- Laboratory of Molecular Neurobiology, Nencki Institute of Experimental Biology of the Polish Academy of Sciences, Warsaw, Poland.
| | - Stefan Loska
- Laboratory of Molecular Neurobiology, Nencki Institute of Experimental Biology of the Polish Academy of Sciences, Warsaw, Poland.
| | - Dominik Cysewski
- Laboratory of RNA Biology and Functional Genomics, Institute of Biochemistry and Biophysics, Warsaw, Poland.
| | - Michal Dabrowski
- Laboratory of Molecular Neurobiology, Nencki Institute of Experimental Biology of the Polish Academy of Sciences, Warsaw, Poland.
| | - Bozena Kaminska
- Laboratory of Molecular Neurobiology, Nencki Institute of Experimental Biology of the Polish Academy of Sciences, Warsaw, Poland.
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Dos Santos PF, Mansur DS. Beyond ISGlylation: Functions of Free Intracellular and Extracellular ISG15. J Interferon Cytokine Res 2017; 37:246-253. [PMID: 28467275 DOI: 10.1089/jir.2016.0103] [Citation(s) in RCA: 67] [Impact Index Per Article: 8.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/04/2023] Open
Abstract
ISG15 is a ubiquitin-like type I IFN-stimulated protein of 15 kDa and is one of the most prominently expressed proteins in viral infections. ISG15 is widely known to be involved in a process called ISGylation, where it binds to over 150 targets from a variety of classes of proteins including central immune signaling pathways such as those mediated by NFκB, JNK, and IRF-3. However, ISG15 also exists in a free form that can act intra- or extracellularly. In vitro and in vivo evidences suggest that free ISG15 play different roles in several cellular processes, from cancer and defense against viral infections to activation of immune cells such as lymphocytes, monocytes, and NK cells. This review discusses the roles of free intracellular and secreted ISG15 approaching questions yet to be answered about the mechanism of action of this protein.
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Affiliation(s)
- Paula Fernandes Dos Santos
- Departament of Microbiology, Immunology and Parasitology, Centro de Ciências Biológicas, Universidade Federal de Santa Catarina , Santa Catarina, Brazil
| | - Daniel Santos Mansur
- Departament of Microbiology, Immunology and Parasitology, Centro de Ciências Biológicas, Universidade Federal de Santa Catarina , Santa Catarina, Brazil
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34
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Hermann M, Bogunovic D. ISG15: In Sickness and in Health. Trends Immunol 2017; 38:79-93. [PMID: 27887993 DOI: 10.1016/j.it.2016.11.001] [Citation(s) in RCA: 79] [Impact Index Per Article: 9.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2016] [Revised: 10/31/2016] [Accepted: 11/01/2016] [Indexed: 12/11/2022]
Abstract
ISG15 is a type I interferon (IFN)-inducible gene encoding a protein with pleiotropic functions, acting both as a soluble molecule and as a protein modifier. Surprisingly, and despite the antiviral functions of ISG15 described in mice, humans born with inactivating mutations of ISG15 do not present with any overt viral phenotype, but are highly susceptible to environmental mycobacteria and have autoinflammatory disease presentations. In vitro, ISG15 deficiency also leads to persistently high levels of type I IFN-stimulated gene expression and to increased resistance to all viruses tested to date. This suggests that ISG15 deficiency increases antiviral responses in humans, in stark contrast to expectations based on mouse experiments. We discuss here the roles of each of the forms of ISG15 in health and disease, as well as the differences between species.
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Affiliation(s)
- Mark Hermann
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, NY, NY 10029, USA
| | - Dusan Bogunovic
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, NY, NY 10029, USA; Department of Pediatrics, Icahn School of Medicine at Mount Sinai, NY, NY 10029, USA; The Mindich Child Health and Development Institute, Icahn School of Medicine at Mount Sinai, NY, NY 10029, USA.
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35
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Jeon YJ, Park JH, Chung CH. Interferon-Stimulated Gene 15 in the Control of Cellular Responses to Genotoxic Stress. Mol Cells 2017; 40:83-89. [PMID: 28241406 PMCID: PMC5339507 DOI: 10.14348/molcells.2017.0027] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/21/2017] [Accepted: 02/23/2017] [Indexed: 12/15/2022] Open
Abstract
Error-free replication and repair of DNA are pivotal to organisms for faithful transmission of their genetic information. Cells orchestrate complex signaling networks that sense and resolve DNA damage. Post-translational protein modifications by ubiquitin and ubiquitin-like proteins, including SUMO and NEDD8, are critically involved in DNA damage response (DDR) and DNA damage tolerance (DDT). The expression of interferon-stimulated gene 15 (ISG15), the first identified ubiquitin-like protein, has recently been shown to be induced under various DNA damage conditions, such as exposure to UV, camptothecin, and doxorubicin. Here we overview the recent findings on the role of ISG15 and its conjugation to target proteins (e.g., p53, ΔNp63α, and PCNA) in the control of cellular responses to genotoxic stress, such as the inhibition of cell growth and tumorigenesis.
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Affiliation(s)
- Young Joo Jeon
- Department of Biochemistry, Chungnam National University School of Medicine, Daejeon 35015,
Korea
- Department of Medical Science, Chungnam National University School of Medicine, Daejeon 35015,
Korea
| | - Jong Ho Park
- School of Biological Sciences, College of Natural Sciences, Seoul National University, Seoul 08826,
Korea
| | - Chin Ha Chung
- School of Biological Sciences, College of Natural Sciences, Seoul National University, Seoul 08826,
Korea
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36
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Villarroya-Beltri C, Baixauli F, Mittelbrunn M, Fernández-Delgado I, Torralba D, Moreno-Gonzalo O, Baldanta S, Enrich C, Guerra S, Sánchez-Madrid F. ISGylation controls exosome secretion by promoting lysosomal degradation of MVB proteins. Nat Commun 2016; 7:13588. [PMID: 27882925 PMCID: PMC5123068 DOI: 10.1038/ncomms13588] [Citation(s) in RCA: 352] [Impact Index Per Article: 39.1] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/24/2015] [Accepted: 10/18/2016] [Indexed: 12/30/2022] Open
Abstract
Exosomes are vesicles secreted to the extracellular environment through fusion with the plasma membrane of specific endosomes called multivesicular bodies (MVB) and mediate cell-to-cell communication in many biological processes. Posttranslational modifications are involved in the sorting of specific proteins into exosomes. Here we identify ISGylation as a ubiquitin-like modification that controls exosome release. ISGylation induction decreases MVB numbers and impairs exosome secretion. Using ISG15-knockout mice and mice expressing the enzymatically inactive form of the de-ISGylase USP18, we demonstrate in vitro and in vivo that ISG15 conjugation regulates exosome secretion. ISG15 conjugation triggers MVB co-localization with lysosomes and promotes the aggregation and degradation of MVB proteins. Accordingly, inhibition of lysosomal function or autophagy restores exosome secretion. Specifically, ISGylation of the MVB protein TSG101 induces its aggregation and degradation, being sufficient to impair exosome secretion. These results identify ISGylation as a novel ubiquitin-like modifier in the control of exosome production. Multivesicular bodies (MVB) are endosomal compartments that can either fuse with the plasma membrane for the secretion of exosomes, or fuse with the lysosome and be degraded along with their contents. Here, the authors show that ISGylation of the MVB protein TSG101 impairs exosome secretion and acts as a regulator of MVB fate.
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Affiliation(s)
- Carolina Villarroya-Beltri
- Centro Nacional de Investigaciones Cardiovasculares (CNIC), 28029 Madrid, Spain.,Immunology Service, Hospital de la Princesa, Instituto Investigación Sanitaria Princesa, Universidad Autónoma de Madrid, 28006 Madrid, Spain
| | - Francesc Baixauli
- Centro Nacional de Investigaciones Cardiovasculares (CNIC), 28029 Madrid, Spain.,Immunology Service, Hospital de la Princesa, Instituto Investigación Sanitaria Princesa, Universidad Autónoma de Madrid, 28006 Madrid, Spain
| | - María Mittelbrunn
- Centro Nacional de Investigaciones Cardiovasculares (CNIC), 28029 Madrid, Spain
| | - Irene Fernández-Delgado
- Centro Nacional de Investigaciones Cardiovasculares (CNIC), 28029 Madrid, Spain.,Immunology Service, Hospital de la Princesa, Instituto Investigación Sanitaria Princesa, Universidad Autónoma de Madrid, 28006 Madrid, Spain
| | - Daniel Torralba
- Centro Nacional de Investigaciones Cardiovasculares (CNIC), 28029 Madrid, Spain.,Immunology Service, Hospital de la Princesa, Instituto Investigación Sanitaria Princesa, Universidad Autónoma de Madrid, 28006 Madrid, Spain
| | - Olga Moreno-Gonzalo
- Centro Nacional de Investigaciones Cardiovasculares (CNIC), 28029 Madrid, Spain.,Immunology Service, Hospital de la Princesa, Instituto Investigación Sanitaria Princesa, Universidad Autónoma de Madrid, 28006 Madrid, Spain
| | - Sara Baldanta
- Department of Preventive Medicine, Public Health and Microbiology, Universidad Autónoma de Madrid, 28029 Madrid, Spain
| | - Carlos Enrich
- Departament de Biomedicina, Unitat de Biologia Cel·lular, Centre de Recerca Biomèdica CELLEX, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Facultat de Medicina, Universitat de Barcelona, 08036 Barcelona, Spain
| | - Susana Guerra
- Department of Preventive Medicine, Public Health and Microbiology, Universidad Autónoma de Madrid, 28029 Madrid, Spain
| | - Francisco Sánchez-Madrid
- Centro Nacional de Investigaciones Cardiovasculares (CNIC), 28029 Madrid, Spain.,Immunology Service, Hospital de la Princesa, Instituto Investigación Sanitaria Princesa, Universidad Autónoma de Madrid, 28006 Madrid, Spain
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Kim YJ, Kim ET, Kim YE, Lee MK, Kwon KM, Kim KI, Stamminger T, Ahn JH. Consecutive Inhibition of ISG15 Expression and ISGylation by Cytomegalovirus Regulators. PLoS Pathog 2016; 12:e1005850. [PMID: 27564865 PMCID: PMC5001722 DOI: 10.1371/journal.ppat.1005850] [Citation(s) in RCA: 50] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/11/2016] [Accepted: 08/08/2016] [Indexed: 11/18/2022] Open
Abstract
Interferon-stimulated gene 15 (ISG15) encodes an ubiquitin-like protein that covalently conjugates protein. Protein modification by ISG15 (ISGylation) is known to inhibit the replication of many viruses. However, studies on the viral targets and viral strategies to regulate ISGylation-mediated antiviral responses are limited. In this study, we show that human cytomegalovirus (HCMV) replication is inhibited by ISGylation, but the virus has evolved multiple countermeasures. HCMV-induced ISG15 expression was mitigated by IE1, a viral inhibitor of interferon signaling, however, ISGylation was still strongly upregulated during virus infection. RNA interference of UBE1L (E1), UbcH8 (E2), Herc5 (E3), and UBP43 (ISG15 protease) revealed that ISGylation inhibits HCMV growth by downregulating viral gene expression and virion release in a manner that is more prominent at low multiplicity of infection. A viral regulator pUL26 was found to interact with ISG15, UBE1L, and Herc5, and be ISGylated. ISGylation of pUL26 regulated its stability and inhibited its activities to suppress NF-κB signaling and complement the growth of UL26-null mutant virus. Moreover, pUL26 reciprocally suppressed virus-induced ISGylation independent of its own ISGylation. Consistently, ISGylation was more pronounced in infections with the UL26-deleted mutant virus, whose growth was more sensitive to IFNβ treatment than that of the wild-type virus. Therefore, pUL26 is a viral ISG15 target that also counteracts ISGylation. Our results demonstrate that ISGylation inhibits HCMV growth at multiple steps and that HCMV has evolved countermeasures to suppress ISG15 transcription and protein ISGylation, highlighting the importance of the interplay between virus and ISGylation in productive viral infection. Type I IFN response is a front-line defense against virus infection. Activation of type I IFN signaling leads to expression of a subset of cellular proteins encoded by interferon-stimulated genes (ISGs). ISG15 encodes an ubiquitin-like protein that is covalently conjugated to protein lysine residues. ISG15 modification (ISGylation) of a protein causes changes of protein function. ISGylation is known to inhibit the replication of many viruses, although pro-viral effects of ISGylation are also reported. Given that ISG15 and the enzymes involved in ISGylation are strongly induced upon virus infection, understanding the interplay between virus and ISGylation is an important issue in virus-host interaction. Nevertheless, viral substrates of ISG15 and viral strategies to regulate ISGylation-mediated antiviral responses are limited to only a few examples. In this study we demonstrate that ISGylation suppresses human cytomegalovirus (HCMV) infection but the virus is armed with countermeasures that consecutively reduce ISG15 transcription and protein ISGylation. Interestingly, a viral ISG15 target is found to inhibit ISGylation. This study highlights that ISGylation is a critical innate immune response against HCMV infection and interfering with ISG15-mediated anti-viral immunity is critical for productive viral infection.
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Affiliation(s)
- Ye Ji Kim
- Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon, Republic of Korea
| | - Eui Tae Kim
- Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon, Republic of Korea
| | - Young-Eui Kim
- Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon, Republic of Korea
| | - Myoung Kyu Lee
- Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon, Republic of Korea
| | - Ki Mun Kwon
- Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon, Republic of Korea
| | - Keun Il Kim
- Department of Biological Sciences, Sookmyung Women's University, Seoul, Republic of Korea
| | - Thomas Stamminger
- Institute for Clinical and Molecular Virology, University of Erlangen-Nuremberg, Schlossgarten, Erlangen, Germany
| | - Jin-Hyun Ahn
- Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon, Republic of Korea
- * E-mail:
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Luo H. Interplay between the virus and the ubiquitin-proteasome system: molecular mechanism of viral pathogenesis. Curr Opin Virol 2015; 17:1-10. [PMID: 26426962 PMCID: PMC7102833 DOI: 10.1016/j.coviro.2015.09.005] [Citation(s) in RCA: 118] [Impact Index Per Article: 11.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/13/2015] [Revised: 09/08/2015] [Accepted: 09/15/2015] [Indexed: 01/24/2023]
Abstract
Many viruses have evolved to utilize the host UPS for their own benefit. Viruses subvert the UPS to maintain optimal level/function of viral proteins. Viruses exploit the UPS to degrade host proteins which impede viral growth. The UPS serves as an important host anti-viral defense mechanism. The UPS is inhibited by some viruses to prevent viral clearance. The ubiquitin–proteasome system (UPS) plays a central role in a wide range of fundamental cellular functions by ensuring protein quality control and through maintaining a critical level of important regulatory proteins. Viruses subvert or manipulate this cellular machinery to favor viral propagation and to evade host immune response. The UPS serves as a double-edged sword in viral pathogenesis: on the one hand, the UPS is utilized by many viruses to maintain proper function and level of viral proteins; while on the other hand, the UPS constitutes a host defense mechanism to eliminate viral components. To combat this host anti-viral machinery, viruses have evolved to employ the UPS to degrade or inactivate cellular proteins that limit viral growth. This review will highlight our current knowledge pertaining to the different roles for the UPS in viral pathogenesis.
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Affiliation(s)
- Honglin Luo
- Centre for Heart Lung Innovation, St. Paul's Hospital, Vancouver, BC, Canada; Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, BC, Canada.
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Mechanism of Dose-Dependent Regulation of UBE1L by Polyphenols in Human Bronchial Epithelial Cells. J Cell Biochem 2015; 116:1553-62. [DOI: 10.1002/jcb.25109] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/10/2014] [Accepted: 01/23/2015] [Indexed: 11/07/2022]
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40
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Selective inactivation of USP18 isopeptidase activity in vivo enhances ISG15 conjugation and viral resistance. Proc Natl Acad Sci U S A 2015; 112:1577-82. [PMID: 25605921 DOI: 10.1073/pnas.1412881112] [Citation(s) in RCA: 94] [Impact Index Per Article: 9.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
Protein modification by the ubiquitin-like protein ISG15 is an interferon (IFN) effector system, which plays a major role in antiviral defense. ISG15 modification is counteracted by the isopeptidase USP18, a major negative regulator of IFN signaling, which was also shown to exert its regulatory function in an isopeptidase-independent manner. To dissect enzymatic and nonenzymatic functions of USP18 in vivo, we generated knock-in mice (USP18(C61A/C61A)) expressing enzymatically inactive USP18. USP18(C61A/C61A) mice displayed increased levels of ISG15 conjugates, validating that USP18 is a major ISG15 isopeptidase in vivo. Unlike USP18(-/-) mice, USP18(C61A/C61A) animals did not exhibit morphological abnormalities, fatal IFN hypersensitivity, or increased lethality, clearly showing that major USP18 functions are unrelated to its protease activity. Strikingly, elevated ISGylation in USP18(C61A/C61A) mice was accompanied by increased viral resistance against vaccinia virus and influenza B virus infections. Enhanced resistance upon influenza B infection in USP18(C61A/C61A) mice was completely reversed in USP18(C61A/C61A) mice, which additionally lack ISG15, providing evidence that the observed reduction in viral titers is ISG15 dependent. These results suggest that increasing ISGylation by specific inhibition of USP18 protease activity could constitute a promising antiviral strategy with only a minimal risk of severe adverse effects.
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Protein interferon-stimulated gene 15 conjugation delays but does not overcome coronavirus proliferation in a model of fulminant hepatitis. J Virol 2014; 88:6195-204. [PMID: 24648452 DOI: 10.1128/jvi.03801-13] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022] Open
Abstract
UNLABELLED Coronaviruses express a deubiquitinating protein, the papain-like protease-2 (PLP2), that removes both ubiquitin and the ubiquitin-like interferon (IFN)-stimulated gene 15 (ISG15) protein from target proteins. ISG15 has antiviral activity against a number of viruses; therefore, we examined the effect of ISG15 conjugation (ISGylation) in a model of acute viral hepatitis induced by the murine hepatitis virus strain 3 (MHV-3) coronavirus. Mice deficient in the ISG15 deconjugating enzyme, ubiquitin-specific peptidase-18 (USP18), accumulate high levels of ISG15-conjugated proteins and are hypersensitive to type I IFN. Infecting USP18(-/-) mice with MHV-3 resulted in extended survival (8 ± 1.2 versus 4 days) and in improved liver histology, a decreased inflammatory response, and viral titers 1 to 2 logs lower than in USP18(+/+) mice. The suppression of viral replication was not due to increased IFN since infected USP18(-/-) mice had neither increased hepatic IFN-α, -β, or -γ mRNA nor circulating protein. Instead, delayed MHV-3 replication coincided with high levels of cellular ISGylation. Decreasing ISGylation by knockdown of the ISG15 E1 enzyme, Ube1L, in primary USP18(+/+) and USP18(-/-) hepatocytes led to increased MHV-3 replication. Both in vitro and in vivo, increasing MHV-3 titers were coincident with increased PLP2 mRNA and decreased ISGylation over the course of infection. The pharmacologic inhibition of the PLP2 enzyme in vitro led to decreased MHV-3 replication. Overall, these results demonstrate the antiviral effect of ISGylation in an in vivo model of coronavirus-induced mouse hepatitis and illustrate that PLP2 manipulates the host innate immune response through the ISG15/USP18 pathway. IMPORTANCE There have been a number of serious worldwide pandemics due to widespread infections by coronavirus. This virus (in its many forms) is difficult to treat, in part because it is very good at finding "holes" in the way that the host (the infected individual) tries to control and eliminate the virus. In this study, we demonstrate that an important host viral defense-the ISG15 pathway-is only partially effective in controlling severe coronavirus infection. Activation of the pathway is very good at suppressing viral production, but over time the virus overwhelms the host response and the effects of the ISG15 pathway. These data provide insight into host-virus interactions during coronavirus infection and suggest that the ISG15 pathway is a reasonable target for controlling severe coronavirus infection although the best treatment will likely involve multiple pathways and targets.
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Liu K, Liao X, Zhou B, Yao H, Fan S, Chen P, Miao D. Porcine alpha interferon inhibit Japanese encephalitis virus replication by different ISGs in vitro. Res Vet Sci 2013; 95:950-6. [DOI: 10.1016/j.rvsc.2013.08.008] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/16/2013] [Revised: 08/01/2013] [Accepted: 08/12/2013] [Indexed: 10/26/2022]
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Yángüez E, García-Culebras A, Frau A, Llompart C, Knobeloch KP, Gutierrez-Erlandsson S, García-Sastre A, Esteban M, Nieto A, Guerra S. ISG15 regulates peritoneal macrophages functionality against viral infection. PLoS Pathog 2013; 9:e1003632. [PMID: 24137104 PMCID: PMC3796851 DOI: 10.1371/journal.ppat.1003632] [Citation(s) in RCA: 32] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2013] [Accepted: 07/29/2013] [Indexed: 12/24/2022] Open
Abstract
Upon viral infection, the production of type I interferon (IFN) and the subsequent upregulation of IFN stimulated genes (ISGs) generate an antiviral state with an important role in the activation of innate and adaptive host immune responses. The ubiquitin-like protein (UBL) ISG15 is a critical IFN-induced antiviral molecule that protects against several viral infections, but the mechanism by which ISG15 exerts its antiviral function is not completely understood. Here, we report that ISG15 plays an important role in the regulation of macrophage responses. ISG15−/− macrophages display reduced activation, phagocytic capacity and programmed cell death activation in response to vaccinia virus (VACV) infection. Moreover, peritoneal macrophages from mice lacking ISG15 are neither able to phagocyte infected cells nor to block viral infection in co-culture experiments with VACV-infected murine embryonic fibroblast (MEFs). This phenotype is independent of cytokine production and secretion, but clearly correlates with impaired activation of the protein kinase AKT in ISG15 knock-out (KO) macrophages. Altogether, these results indicate an essential role of ISG15 in the cellular immune antiviral response and point out that a better understanding of the antiviral responses triggered by ISG15 may lead to the development of therapies against important human pathogens. Modification of proteins by ubiquitin (UB) and ubiquitin-like proteins (UBLs) are key regulatory processes of the innate and adaptive immune response. Interferon (IFN) stimulated gene product 15 (ISG15) is an ubiquitin-like protein modifier, which is reversibly conjugated to different viral and cellular proteins mediating considerable antiviral responses. In turn, many viruses, including poxviruses, have evolved strategies to block the antiviral and inflammatory effects of the innate immune responses to keep cells alive until virus replication is completed. Here, we describe a novel function of ISG15 in the control of macrophages activation, phagocytosis and apoptosis in response to viral infection. These processes are essential for the self-defense mechanism to protect animals from infectious disease and could be crucial to understand the ISG15 antiviral activity described in animal models.
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Affiliation(s)
- Emilio Yángüez
- Institute of Medical Virology, University of Zurich, Zurich, Switzerland
| | - Alicia García-Culebras
- Department of Preventive Medicine and Public Health, Universidad Autónoma, Madrid, Spain
| | - Aldo Frau
- Department of Preventive Medicine and Public Health, Universidad Autónoma, Madrid, Spain
| | - Catalina Llompart
- Department of Molecular and Cellular Biology, Centro Nacional de Biotecnología CSIC, Madrid, Spain
- Ciber de Enfermedades Respiratorias, Madrid, Spain
| | | | | | - Adolfo García-Sastre
- Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America
- Department of Medicine, Division of Infectious Diseases, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America
- Global Health and Emerging Pathogens Institute, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America
| | - Mariano Esteban
- Department of Molecular and Cellular Biology, Centro Nacional de Biotecnología CSIC, Madrid, Spain
| | - Amelia Nieto
- Department of Molecular and Cellular Biology, Centro Nacional de Biotecnología CSIC, Madrid, Spain
- Ciber de Enfermedades Respiratorias, Madrid, Spain
| | - Susana Guerra
- Department of Preventive Medicine and Public Health, Universidad Autónoma, Madrid, Spain
- * E-mail:
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Morales DJ, Lenschow DJ. The antiviral activities of ISG15. J Mol Biol 2013; 425:4995-5008. [PMID: 24095857 PMCID: PMC4090058 DOI: 10.1016/j.jmb.2013.09.041] [Citation(s) in RCA: 195] [Impact Index Per Article: 16.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2013] [Revised: 09/27/2013] [Accepted: 09/30/2013] [Indexed: 01/01/2023]
Abstract
Post-translational protein modification is an important strategy for the regulation of the cell proteome independent of the need for new gene expression. Ubiquitin and ubiquitin-like modifiers mediate the regulation of protein levels, signaling pathways, vesicular trafficking, and many other cellular processes through their covalent conjugation to proteins. Interferon stimulated gene 15 (ISG15) is a ubiquitin-like modifier induced by type I interferon. In addition to conjugating to potentially hundreds of target proteins, ISG15 can be found in an unconjugated form both inside of the cell and released from interferon stimulated cells into the extracellular environment. Due to its robust expression after type I interferon stimulation and the broad panel of proteins that it targets, ISG15 has drawn much attention as a potential regulator of the immune response and has been shown to mediate protection in a number of different viral infection models. Here we will review the current state of the field of ISG15, the viruses against which ISG15 mediates protection, and the mechanisms by which ISG15 exerts antiviral activity.
ISG15 is an interferon-induced ubiquitin-like modifier that plays an important role during host responses to viral infections. ISG15 mediates these functions in a conjugation-dependent manner by targeting both host and viral proteins. Unconjugated ISG15 can also regulate the host response to viral infection through distinct mechanisms of action.
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Affiliation(s)
- David J Morales
- Department of Medicine and Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110, USA
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Campbell JA, Lenschow DJ. Emerging roles for immunomodulatory functions of free ISG15. J Interferon Cytokine Res 2013; 33:728-38. [PMID: 24010825 DOI: 10.1089/jir.2013.0064] [Citation(s) in RCA: 33] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Abstract
Type I interferons (IFNs) exert their effects through the induction of hundreds of IFN-stimulated genes (ISGs), many of which function by inhibiting viral replication and modulating immune responses. ISG15, a di-ubiquitin-like protein, is one of the most abundantly induced ISGs and is critical for control of certain viral and bacterial infections. Like ubiquitin, ISG15 is covalently conjugated to target proteins. In addition, free unconjugated ISG15 is present both intra- and extracellularly. Although much remains to be learned about conjugated ISG15, even less is known about the 2 free forms of ISG15. This article focuses on the role that ISG15 plays during the host response to pathogen challenge, in particular on the recent observations describing the immunomodulatory properties of free ISG15 and its potential implication in disease pathogenesis.
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Affiliation(s)
- Jessica A Campbell
- Department of Internal Medicine, Washington University School of Medicine , St. Louis, Missouri
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Zebrafish ISG15 exerts a strong antiviral activity against RNA and DNA viruses and regulates the interferon response. J Virol 2013; 87:10025-36. [PMID: 23824820 DOI: 10.1128/jvi.01294-12] [Citation(s) in RCA: 74] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
ISG15, a 15-kDa interferon-induced protein that participates in antiviral defenses of mammals, is highly conserved among vertebrates. In fish, as in mammals, viral infection and interferon treatment induce isg15 expression. The two ubiquitin-like domains of ISG15 and the presence of a consensus LRLRGG sequence in the C-terminal region, which is required for the covalent conjugation to a substrate protein, are also conserved in fish. Our data demonstrate that overexpression of zebrafish ISG15 (zf-ISG15) in EPC cells is sufficient to inhibit viral infection by RNA viruses belonging to the genera Novirhabdovirus and Birnavirus and by DNA viruses of the genus Iridovirus. In coexpression experiments with IHNV proteins, we demonstrate specific ISGylation of phosphoprotein and nonvirion protein. Mutation of the glycine residues in the consensus LRLRGG motif abolishes zf-ISG15 conjugation to these proteins and the cellular protection against viral infection, thus connecting ISGylation and ISG15-dependent viral restriction. Additionally, zf-ISG15 overexpression triggers induction of the rig-I and viperin genes as well as, to a lesser extent, the IFN gene. Overall, our data demonstrate the antiviral effect of a fish ISG15 protein, revealing the conservation among vertebrates of an ISGylation mechanism likely directed against viruses. Furthermore, our findings indicate that zf-ISG15 affects the IFN system at several levels, and its study shall shed further light on the evolution of the complex regulation of the innate antiviral response in vertebrate cells.
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47
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Zhao C, Collins MN, Hsiang TY, Krug RM. Interferon-induced ISG15 pathway: an ongoing virus-host battle. Trends Microbiol 2013; 21:181-6. [PMID: 23414970 PMCID: PMC3622817 DOI: 10.1016/j.tim.2013.01.005] [Citation(s) in RCA: 102] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/24/2012] [Revised: 01/17/2013] [Accepted: 01/18/2013] [Indexed: 12/21/2022]
Abstract
ISG15 is an interferon (IFN)-induced ubiquitin-like protein that is conjugated to target proteins via the sequential action of three enzymes that are also induced by IFN. Unlike ubiquitin, which is highly conserved, the sequence of ISG15 varies between species. ISG15 conjugation inhibits many viruses, and free (unconjugated) ISG15 can also act as an antiviral protein. In this review, we focus on the antiviral role of ISG15 conjugation and on countermeasures employed by several viruses. The countermeasure by influenza B virus is unique in that it exhibits species specificity. Only the antiviral activity of human and non-human primate ISG15s can be blocked, providing one possible explanation for the restriction of influenza B virus to humans.
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Affiliation(s)
- Chen Zhao
- Department of Molecular Genetics and Microbiology, Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, TX 78712, USA
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48
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An attenuating mutation in a neurovirulent Sindbis virus strain interacts with the IPS-1 signaling pathway in vivo. Virology 2012; 435:269-80. [PMID: 23084425 DOI: 10.1016/j.virol.2012.09.008] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/26/2012] [Revised: 08/27/2012] [Accepted: 09/13/2012] [Indexed: 12/24/2022]
Abstract
The AR86 strain of Sindbis virus causes lethal neurologic disease in adult mice. Previous studies have identified a virulence determinant at nonstructural protein (nsP) 1 position 538 that regulates neurovirulence, modulates clearance from the CNS, and interferes with the type I interferon pathway. The studies herein demonstrate that in the absence of type I interferon signaling, the attenuated mutant exhibited equivalent virulence to S300 virus. Furthermore, both S300 and nsP1 T538I viruses displayed similar neurovirulence and replication kinetics in IPS-1-/- mice. TRIF dependent signaling played a modest role in protecting against disease by both S300 and nsP1 T538I, but did not contribute to control of nsP1 T538I replication within the CNS, while MyD88 played no role in the disease process. These results indicate that the control of the nsP1 T538I mutant virus is largely mediated by IPS-1-dependent RLR signaling, with TRIF-dependent TLR signaling also contributing to protection from virus-induced neurologic disease.
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49
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Choi AG, Wong J, Marchant D, Luo H. The ubiquitin-proteasome system in positive-strand RNA virus infection. Rev Med Virol 2012; 23:85-96. [PMID: 22782620 PMCID: PMC7169083 DOI: 10.1002/rmv.1725] [Citation(s) in RCA: 31] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2012] [Revised: 05/29/2012] [Accepted: 06/18/2012] [Indexed: 12/12/2022]
Abstract
Positive-stranded RNA viruses, like many other viruses, have evolved to exploit the host cellular machinery to their own advantage. In eukaryotic cells, the ubiquitin-proteasome system (UPS) that serves as the major intracellular pathway for protein degradation and modification plays a crucial role in the regulation of many fundamental cellular functions. A growing amount of evidence has suggested that the UPS can be utilized by positive-sense RNA viruses. The UPS eliminates excess viral proteins that prevent viral replication and modulates the function of viral proteins through post-translational modification mediated by ubiquitin or ubiquitin-like proteins. This review will discuss the current understanding of how positive RNA viruses have evolved various mechanisms to usurp the host UPS to modulate the function and stability of viral proteins. In addition to the pro-viral function, UPS-mediated viral protein degradation may also constitute a host defense process against some positive-stranded RNA viral infections. This issue will also be discussed in the current review.
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Affiliation(s)
- Alex GoEun Choi
- UBC James Hogg Research Centre, Institute for Heart + Lung Health, St. Paul's Hospital, Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, BC, Canada
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50
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Budding of Enveloped Viruses: Interferon-Induced ISG15-Antivirus Mechanisms Targeting the Release Process. Adv Virol 2012; 2012:532723. [PMID: 22666250 PMCID: PMC3362814 DOI: 10.1155/2012/532723] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2012] [Accepted: 03/12/2012] [Indexed: 11/17/2022] Open
Abstract
Pathogenic strains of viruses that infect humans are encapsulated in membranes derived from the host cell in which they infect. After replication, these viruses are released by a budding process that requires cell/viral membrane scission. As such, this represents a natural target for innate immunity mechanisms to interdict enveloped virus spread and recent advances in this field will be the subject of this paper.
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