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Human adenovirus – host cell interplay: The role of the cellular zinc finger proteins and mitochondrial DNA
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Human adenovirus (HAdV) is an abundant DNA virus with significant clinical relevance since it cauces a variety of respiratory, ocular, and gastrointestinal diseases. It is also intensively used as a therapeutic tool to treat cancers and to boost immune responses. In order to achieve a better control over the HAdV epidemiology and improved utilization for clinical applications, it is crucial to understand the molecular interaction between the host cell and HAdV.

The aim of the current thesis is to delineate the molecular interactions between HAdV type 5 (HAdV-C5) protein VII (pVII), two cellular zinc finger proteins (MKRN1, ZNF622)  and mitochondrial DNA (mtDNA). In paper I, we have identified MKRN1 as one of the novel pVII-interacting proteins. Surprisingly, endogenous MKRN1 protein is down-regulated in the HAdV-infected cells due to its proteasomal degradation. Further, the pVII(wt) promoted  MKRN1 self-ubiquitination, which may explain the overall instability of the MKRN1 protein in the infected cells. In addition, we show that the MKRN1 protein is also down-regulated in measles virus- and vesicular stomatitis virus-infected cells. In paper II, we report that the cellular ZNF622 protein interacts with the pVII protein. Intriguingly, ZNF622 expression was enhanced in HAdV-C5-infected cells, implying its anti-viral role. Surprisingly, lack of the ZNF622 protein significantly enhanced formation of the infectious HAdV-C5 virions. Finally, we propose a model how the ZNF622/NPM1/pVII protein complex regulates the pVII protein binding to viral DNA. In paper III, we report that HAdV-C5 infection enhanced mtDNA release into cytosol. The enhanced mtDNA release can be partially explained by accumulation of the pVII protein since its down-regulation diminished mtDNA release into cyotosol. We also report pVII-regulated gene expression profile and show that cellular cytokine IL-32 mRNA accumulates in response to the pVII protein expression.

Collectively, in this thesis we provide molecular characterization how two cellular zinc finger proteins (MKRN1 and ZNF622) and mtDNA behave in the context of lytic HAdV-C5 infection. The ZNF622 may act as a bona fide anti-viral factor blocking infectious virion formation via targeting the essential viral core protein pVII. The MKRN1 protein is efficiently eliminated in the infected cells, highlighting the essence of HAdV-C5-controlled proteasome. Finally, dynamical change of mtDNA induced by HAdV-C5 infection, might initiate a novel signaling pathway beneficial for the cells or the viruses.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2019. , p. 61
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1583
Keywords [en]
Adenovirus, Protein VII, Ubiquitination, Chromatin remodeling, Mitochondria
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:uu:diva-389805ISBN: 978-91-513-0696-4 (print)OAI: oai:DiVA.org:uu-389805DiVA, id: diva2:1339340
Public defence
2019-09-05, C8:305, Biomedical Centrum (BMC), Husargatan 3, Uppsala, 09:00 (English)
Opponent
Supervisors
Available from: 2019-08-15 Created: 2019-07-29 Last updated: 2019-09-17
List of papers
1. Human Adenovirus Infection Causes Cellular E3 Ubiquitin Ligase MKRN1 Degradation Involving the Viral Core Protein pVII.
Open this publication in new window or tab >>Human Adenovirus Infection Causes Cellular E3 Ubiquitin Ligase MKRN1 Degradation Involving the Viral Core Protein pVII.
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2018 (English)In: Journal of Virology, ISSN 0022-538X, E-ISSN 1098-5514, Vol. 92, no 3, article id e01154-17Article in journal (Refereed) Published
Abstract [en]

Human adenoviruses (HAdVs) are common human pathogens encoding a highly abundant histone-like core protein, VII, which is involved in nuclear delivery and protection of viral DNA as well as in sequestering immune danger signals in infected cells. The molecular details of how protein VII acts as a multifunctional protein have remained to a large extent enigmatic. Here we report the identification of several cellular proteins interacting with the precursor pVII protein. We show that the cellular E3 ubiquitin ligase MKRN1 is a novel precursor pVII-interacting protein in HAdV-C5-infected cells. Surprisingly, the endogenous MKRN1 protein underwent proteasomal degradation during the late phase of HAdV-C5 infection in various human cell lines. MKRN1 protein degradation occurred independently of the HAdV E1B55K and E4orf6 proteins. We provide experimental evidence that the precursor pVII protein binding enhances MKRN1 self-ubiquitination, whereas the processed mature VII protein is deficient in this function. Based on these data, we propose that the pVII protein binding promotes MKRN1 self-ubiquitination, followed by proteasomal degradation of the MKRN1 protein, in HAdV-C5-infected cells. In addition, we show that measles virus and vesicular stomatitis virus infections reduce the MKRN1 protein accumulation in the recipient cells. Taken together, our results expand the functional repertoire of the HAdV-C5 precursor pVII protein in lytic virus infection and highlight MKRN1 as a potential common target during different virus infections. IMPORTANCE: Human adenoviruses (HAdVs) are common pathogens causing a wide range of diseases. To achieve pathogenicity, HAdVs have to counteract a variety of host cell antiviral defense systems, which would otherwise hamper virus replication. In this study, we show that the HAdV-C5 histone-like core protein pVII binds to and promotes self-ubiquitination of a cellular E3 ubiquitin ligase named MKRN1. This mutual interaction between the pVII and MKRN1 proteins may prime MKRN1 for proteasomal degradation, because the MKRN1 protein is efficiently degraded during the late phase of HAdV-C5 infection. Since MKRN1 protein accumulation is also reduced in measles virus- and vesicular stomatitis virus-infected cells, our results signify the general strategy of viruses to target MKRN1.

Keywords
adenoviruses, proteasome, ubiquitination
National Category
Microbiology
Identifiers
urn:nbn:se:uu:diva-339568 (URN)10.1128/JVI.01154-17 (DOI)000423571600001 ()29142133 (PubMedID)
Funder
Åke Wiberg Foundation, M14-0155Swedish Cancer Society, CAN 2013/350Swedish Research Council, 2006-5038-36531-16Science for Life Laboratory - a national resource center for high-throughput molecular bioscience
Available from: 2018-01-20 Created: 2018-01-20 Last updated: 2019-07-29Bibliographically approved
2. Cellular Zinc Finger Protein 622 Hinders Human Adenovirus Lytic Growth and Limits Binding of the Viral pVII Protein to Virus DNA
Open this publication in new window or tab >>Cellular Zinc Finger Protein 622 Hinders Human Adenovirus Lytic Growth and Limits Binding of the Viral pVII Protein to Virus DNA
2019 (English)In: Journal of Virology, ISSN 0022-538X, E-ISSN 1098-5514, Vol. 93, no 3, article id e01628-18Article in journal (Refereed) Published
Abstract [en]

Human adenovirus (HAdV) encodes a multifunctional DNA-binding protein pVII, which is involved in virus DNA packaging and extracellular immune signaling regulation. Although the pVII is an essential viral protein, its exact role in the virus life cycle and interplay with cellular proteins have remained to a large extent unclear. We have recently identified the cellular zinc finger protein 622 (ZNF622) as a potential pVII-interacting protein. In this study, we describe the functional consequences of the ZNF622-pVII interplay and the role of ZNF622 in the HAdV life cycle. ZNF622 protein expression increased, and it accumulated similarly to the pVII protein in the nuclei of virus-infected cells. The lack of the ZNF622 protein specifically increased pVII binding to viral DNA in the infected cells and elevated the pVII protein levels in the purified virions. In addition, ZNF622 knockout cells showed an increased cell lysis and enhanced accumulation of the infectious virus particles. Protein interaction studies revealed that ZNF622 forms a trimeric complex with the pVII protein and the cellular histone chaperon protein nucleophosmin 1 (NPM1). The integrity of this complex is important since ZNF622 mutations and NPM1 deficiency changed pVII ability to bind viral DNA. Collectively, our results implicate that ZNF622 may act as a cellular antiviral protein hindering lytic HAdV growth and limiting pVII protein binding to viral DNA.

IMPORTANCE Human adenoviruses (HAdVs) are common human pathogens causing a wide range of acute infections. To counteract viral pathogenicity, cells encode a variety of antiviral proteins and noncoding RNAs to block virus growth. In this study, we show that the cellular zinc finger protein 622 (ZNF622) interacts with an essential HAdV protein known as pVII. This mutual interaction limits pVII binding to viral DNA. Further, ZNF622 has a role in HAdV life cycle since the lack of ZNF622 correlates with increased lysis of the infected cells and accumulation of the infectious virions. Together, our study reveals a novel cellular antiviral protein ZNF622, which may impede lytic HAdV growth.

Keywords
adenoviruses, chromatin remodeling
National Category
Microbiology
Identifiers
urn:nbn:se:uu:diva-375801 (URN)10.1128/JVI.01628-18 (DOI)000456001300019 ()30429337 (PubMedID)
Funder
Swedish Research Council, 2006-5038-36531-16Åke Wiberg Foundation, M14-0155Swedish Cancer Society, CAN 2013/350
Available from: 2019-02-13 Created: 2019-02-13 Last updated: 2019-07-29
3. Human adenovirus core protein pVII influences mitochondrial DNA dynamics and expression of the pro-inflammatory cytokine IL-32
Open this publication in new window or tab >>Human adenovirus core protein pVII influences mitochondrial DNA dynamics and expression of the pro-inflammatory cytokine IL-32
(English)Manuscript (preprint) (Other academic)
National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-389804 (URN)
Available from: 2019-07-29 Created: 2019-07-29 Last updated: 2019-07-29

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