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Human adenovirus infection counteracts the anti-viral activity of the cellular MKRN1 E3 ubiquitin ligase
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
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(English)Article in journal (Refereed) Submitted
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
URN: urn:nbn:se:uu:diva-321638OAI: oai:DiVA.org:uu-321638DiVA: diva2:1094344
Available from: 2017-05-09 Created: 2017-05-09 Last updated: 2017-05-09
In thesis
1. Functional characterization of the human adenovirus pVII protein and non-coding VA RNAI
Open this publication in new window or tab >>Functional characterization of the human adenovirus pVII protein and non-coding VA RNAI
2017 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Human adenovirus (HAdV) is a common pathogen causing a broad spectrum of diseases. HAdV encodes the pVII protein, which is involved in nuclear delivery, protection and expression of viral DNA. To suppress the cellular interferon (IFN) and RNA interference (RNAi) systems, HAdVs encode non-coding virus-associated (VA) RNAs. In this thesis we have investigated the functional significance of the pVII protein and VA RNAI in HAdV-5 infected cells.

We report that the propeptide module is the destabilizing element targeting the precursor pVII protein for proteasomal degradation. We also found that the Cul3-based E3 ubiquitin ligase complex alter the precursor pVII protein stability via binding to the propeptide sequence. In addition, we show that inhibition of the Cul3 protein reduces HAdV-5 E1A gene expression. Collectively, our results suggest a novel function for the pVII propeptide module and involvement of Cul3 in viral E1A gene expression.

Our studies show that the cellular E3 ubiquitin ligase MKRN1 is a novel pVII interacting protein in HAdV-5 infected cells. MKRN1 expression reduced the pVII protein accumulation in virus-infected cells and affected infectious virus formation. Surprisingly, the endogenous MKRN1 protein underwent proteasomal degradation during the prolonged HAdV-5 infection. Furthermore, the precursor pVII protein enhanced MKRN1 self-ubiquitination, suggesting the direct involvement of pVII in the initiation of MKRN1 degradation. Hence, we propose that the MKRN1 is a novel antiviral protein and that HAdV-5 infection counteracts its antiviral activity.

In papers III and IV, we tested the ability of various plant and animal virus encoded RNAi/miRNA and IFN suppressor proteins to functionally substitute for the HAdV-5 VA RNAI. Our results revealed that the Vaccinia virus E3L protein was able to partially substitute for the HAdV-5 VA RNAI functions in virus-infected cells. Interestingly, the E3L protein rescued the translational defect but did not stimulate viral capsid mRNA accumulation observed with VA RNA. Additionally, we show that the HAdV-4 and HAdV-37 VA RNAI are more effective in virus replication compared to HAdV-5 and HAdV-12 VA RNAI. In paper IV, we employed a novel triplex-specific probing assay, based on the intercalating and cleaving agent benzoquinoquinaxline 1,10-phenanthroline (BQQ-OP), to unravel triplex structure formation in 
VA RNAI. The BQQ-OP cleavage of HAdV-4 VA RNAI indicates that a potential 
triplex is formed involving the highly conserved stem 4 of the central domain and side 
stem 7. Further, the integrity of HAdV-4 VA RNAI stem 7 contributes to the virus growth in vivo.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2017. 58 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1339
Keyword
Adenovirus, VA RNA, protein VII, Ubiquitination, proteasome, anti-viral
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy) Cell and Molecular Biology
Research subject
Medical Biochemistry; Microbiology; Medical Virology
Identifiers
urn:nbn:se:uu:diva-321641 (URN)978-91-554-9941-9 (ISBN)
Public defence
2017-09-01, Room C8:305, Biomedical Centrum (BMC), Husargatan 3, Uppsala, 13:15 (English)
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Available from: 2017-06-09 Created: 2017-05-09 Last updated: 2017-08-09

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