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postgraduate thesis: Role of middle-east respiratory syndrome coronavirus membrane and open reading frame 8b proteins in type I interferon antagonism
Title | Role of middle-east respiratory syndrome coronavirus membrane and open reading frame 8b proteins in type I interferon antagonism |
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Authors | |
Advisors | |
Issue Date | 2018 |
Publisher | The University of Hong Kong (Pokfulam, Hong Kong) |
Citation | Wong, L. R. [黃樂賢]. (2018). Role of middle-east respiratory syndrome coronavirus membrane and open reading frame 8b proteins in type I interferon antagonism. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR. |
Abstract | Middle-East respiratory syndrome coronavirus (MERS-CoV) was the most recently identified human coronavirus in 2013 that was known to cause acute severe respiratory distress syndrome. MERS-CoV-infected individuals presented clinical symptoms that resembled those of SARS-CoV-infected patients and MERS was thought to be the second human coronavirus identified with fatal outcomes in infected patients. Phylogenetic analysis revealed bat coronaviruses HKU4 and HKU5 as the most closely related counterparts of MERS-CoV in lineage C of Betacoronaviruses. One of the characteristics of MERS-CoV is the ability to suppress innate immune responses, in particular type I interferon (IFN) responses, which was evident by the significantly reduced type I IFN level in infected individuals. Type I IFNs are a group of crucial cytokines that activate the innate immune system by activating the transcription of various interferon-stimulated genes (ISGs), which are innate immune effectors of paramount importance in establishing antiviral state in host and for viral clearance. Efforts in identifying MERS-CoV viral proteins in suppressing type I IFNs revealed that multiple MERS-CoV viral proteins act on various levels of the innate immunity. In this thesis, I have identified two viral proteins, the structural membrane (M) and the accessory open reading frame 8b (ORF8b) proteins encoded by MERS-CoV genome as type I IFN antagonists that specifically suppressed interferon regulatory factor 3 (IRF3)-mediated type I IFN expression induced by Sendai virus, double-stranded RNA analogue polyinosinic:polycytidylic acid (polyI:C) and dominant active form of retinoic-acid inducible gene-I (RIG-IN), which are potent stimuli activating RNA sensing pathway highly releveant to human coronavirus infection while sparing nuclear factor-κB (NF-κB)-mediated type I IFN expression. In particular, I have shown that MERS-CoV M protein acts upstream of IRF3 by interacting with innate immune adapter TRAF3 and prevents the recruitment of TBK1, which serves as the major activation signal for IRF3. Domain mapping suggested that the N-terminal domain of MERS-CoV M protein alone was sufficient for the immunosuppressive effect, whereas the C-terminal domain was devoid of this activity. Also, I have shown that the ectopic expression of MERS-CoV ORF8b inhibited TBK1 and IRF3 activation, which is essential for type I IFN expression. Further analysis by mass spectrometry revealed heat-shock proteins (HSPs) as binding partners of ORF8b and co-IP confirmed specific interaction between HSP70 and ORF8b. I also identified the role of HSP70 as an enhancer of IKKε activation and that ORF8b interact with HSP70 and HSP70 interact with IKKε. Taken together, I have identified a novel role of HSP70 in enhancing IKKε activation through a yet-to-be-known mechanism and that MERS-CoV ORF8b exploits the enhancing effect of HSP70 towards IKKε activation to achieve its suppression of innate immune antiviral responses through type I IFN antagonism. |
Degree | Doctor of Philosophy |
Subject | MERS (Disease) Coronaviruses |
Dept/Program | Biomedical Sciences |
Persistent Identifier | http://hdl.handle.net/10722/274654 |
DC Field | Value | Language |
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dc.contributor.advisor | Jin, D | - |
dc.contributor.advisor | Kok, KH | - |
dc.contributor.author | Wong, Lok-yin, Roy | - |
dc.contributor.author | 黃樂賢 | - |
dc.date.accessioned | 2019-09-09T07:21:25Z | - |
dc.date.available | 2019-09-09T07:21:25Z | - |
dc.date.issued | 2018 | - |
dc.identifier.citation | Wong, L. R. [黃樂賢]. (2018). Role of middle-east respiratory syndrome coronavirus membrane and open reading frame 8b proteins in type I interferon antagonism. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR. | - |
dc.identifier.uri | http://hdl.handle.net/10722/274654 | - |
dc.description.abstract | Middle-East respiratory syndrome coronavirus (MERS-CoV) was the most recently identified human coronavirus in 2013 that was known to cause acute severe respiratory distress syndrome. MERS-CoV-infected individuals presented clinical symptoms that resembled those of SARS-CoV-infected patients and MERS was thought to be the second human coronavirus identified with fatal outcomes in infected patients. Phylogenetic analysis revealed bat coronaviruses HKU4 and HKU5 as the most closely related counterparts of MERS-CoV in lineage C of Betacoronaviruses. One of the characteristics of MERS-CoV is the ability to suppress innate immune responses, in particular type I interferon (IFN) responses, which was evident by the significantly reduced type I IFN level in infected individuals. Type I IFNs are a group of crucial cytokines that activate the innate immune system by activating the transcription of various interferon-stimulated genes (ISGs), which are innate immune effectors of paramount importance in establishing antiviral state in host and for viral clearance. Efforts in identifying MERS-CoV viral proteins in suppressing type I IFNs revealed that multiple MERS-CoV viral proteins act on various levels of the innate immunity. In this thesis, I have identified two viral proteins, the structural membrane (M) and the accessory open reading frame 8b (ORF8b) proteins encoded by MERS-CoV genome as type I IFN antagonists that specifically suppressed interferon regulatory factor 3 (IRF3)-mediated type I IFN expression induced by Sendai virus, double-stranded RNA analogue polyinosinic:polycytidylic acid (polyI:C) and dominant active form of retinoic-acid inducible gene-I (RIG-IN), which are potent stimuli activating RNA sensing pathway highly releveant to human coronavirus infection while sparing nuclear factor-κB (NF-κB)-mediated type I IFN expression. In particular, I have shown that MERS-CoV M protein acts upstream of IRF3 by interacting with innate immune adapter TRAF3 and prevents the recruitment of TBK1, which serves as the major activation signal for IRF3. Domain mapping suggested that the N-terminal domain of MERS-CoV M protein alone was sufficient for the immunosuppressive effect, whereas the C-terminal domain was devoid of this activity. Also, I have shown that the ectopic expression of MERS-CoV ORF8b inhibited TBK1 and IRF3 activation, which is essential for type I IFN expression. Further analysis by mass spectrometry revealed heat-shock proteins (HSPs) as binding partners of ORF8b and co-IP confirmed specific interaction between HSP70 and ORF8b. I also identified the role of HSP70 as an enhancer of IKKε activation and that ORF8b interact with HSP70 and HSP70 interact with IKKε. Taken together, I have identified a novel role of HSP70 in enhancing IKKε activation through a yet-to-be-known mechanism and that MERS-CoV ORF8b exploits the enhancing effect of HSP70 towards IKKε activation to achieve its suppression of innate immune antiviral responses through type I IFN antagonism. | - |
dc.language | eng | - |
dc.publisher | The University of Hong Kong (Pokfulam, Hong Kong) | - |
dc.relation.ispartof | HKU Theses Online (HKUTO) | - |
dc.rights | The author retains all proprietary rights, (such as patent rights) and the right to use in future works. | - |
dc.rights | This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License. | - |
dc.subject.lcsh | MERS (Disease) | - |
dc.subject.lcsh | Coronaviruses | - |
dc.title | Role of middle-east respiratory syndrome coronavirus membrane and open reading frame 8b proteins in type I interferon antagonism | - |
dc.type | PG_Thesis | - |
dc.description.thesisname | Doctor of Philosophy | - |
dc.description.thesislevel | Doctoral | - |
dc.description.thesisdiscipline | Biomedical Sciences | - |
dc.description.nature | published_or_final_version | - |
dc.identifier.doi | 10.5353/th_991044058291903414 | - |
dc.date.hkucongregation | 2018 | - |
dc.identifier.mmsid | 991044058291903414 | - |