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The adenovirus VA RNA-derived miRNAs are not essential for lytic virus growth in tissue culture cells
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.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
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2013 (English)In: Nucleic Acids Research, ISSN 0305-1048, E-ISSN 1362-4962, Vol. 41, no 9, 4802-4812 p.Article in journal (Refereed) Published
Abstract [en]

At late times during a lytic infection human adenovirus type 5 produces ∼10(8) copies per cell of virus-associated RNA I (VA RNAI). This short highly structured RNA polymerase III transcript has previously been shown to be essential for lytic virus growth. A fraction of VA RNAI is processed by Dicer into small RNAs, so-called mivaRNAIs, which are efficiently incorporated into the RNA-induced silencing complex. Here, we constructed recombinant adenoviruses with mutations in the seed sequence of both the 5'- and the 3'-strand of the mivaRNAI duplex. The results showed that late viral protein synthesis, as well as new virus progeny formation, was essentially unaffected by the seed sequence mutations under lytic replicative conditions in HeLa or HEK293 cells. Collectively, our results suggest that either strand of the mivaRNAI duplex does not have target mRNA interactions that are critical for the establishment of virus growth under lytic conditions. Further, by depletion of protein kinase R (PKR) in HEK293 cells, we show that the suppressive effect of VA RNAI on the interferon-induced PKR pathway is most critical for late gene expression.

Place, publisher, year, edition, pages
2013. Vol. 41, no 9, 4802-4812 p.
National Category
Natural Sciences
URN: urn:nbn:se:uu:diva-198031DOI: 10.1093/nar/gkt172ISI: 000318570000013PubMedID: 23525465OAI: oai:DiVA.org:uu-198031DiVA: diva2:615035
Available from: 2013-04-08 Created: 2013-04-08 Last updated: 2016-06-01Bibliographically approved
In thesis
1. Adenoviral small non-coding RNAs: A Structural and Functional Charaterization
Open this publication in new window or tab >>Adenoviral small non-coding RNAs: A Structural and Functional Charaterization
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Since their discovery in 1953, adenoviruses have significantly contributed to the understanding of virus-host cell interactions, including mechanistic details of cellular processes such as cell cycle control and alternative RNA splicing. Among the first characterized adenoviral genes were the virus-associated RNAs (VA RNAI/II), which are produced in massive amount during a lytic infection. The VA RNAs perform multiple functions and are required for a successful adenovirus life cycle. More recently, it was shown that the VA RNAs are processed into small viral miRNAs, so-called mivaRNAs, which interfere with the function of the cellular RNAi/miRNA machinery.

In papers I and II, we focused on a structural and functional characterization of the mivaRNAs using two approaches. Firstly, we created a model system where the predicted miRNA-like function of mivaRNAI could be investigated, without interfering with other VA RNA functions. This was accomplished by construction of recombinant adenoviruses, in which the seed sequence of mivaRNAI was altered. The results showed that in cell culture experiments the mivaRNAI seed sequence mutants grew as the wild type virus, suggesting that the mivaRNAs are not required during the lytic phase of an adenovirus infection. Secondly, we showed that the VA RNAs from different human adenoviruses (Ad4, Ad5, Ad11 and Ad37) undergo the same type of Dicer-dependent processing into mivaRNAs, which subsequently are loaded onto the RNA induced silencing complex (RISC), albeit with different efficiencies.

In paper III, we demonstrated that the promoter proximal region of the adenovirus major late promoter (MLP) produces a novel non-canonical class of small RNAs, which we termed the MLP-TSS-sRNAs. Surprisingly the MLP-TSS-sRNA maintains the m7G-cap structure while bound to Ago2 containing RISC. These complexes are functional suppressing expression of target mRNAs with complementary binding site. Most importantly, the MLP-TSS-sRNA limits the efficiency of viral DNA replication probably through a targeting of the E2B mRNAs, which are transcribed in the antisense orientation. In conclusion, the MLP-TSS-sRNA represents the first viral small RNA, which has been shown to have a function as a regulator of an adenovirus infection.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2016. 49 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1216
non-coding RNA miRNA mivaRNA canonical Dicer Ago2 TSS m7G Cap Adenovirus replication E2B
National Category
Medical and Health Sciences
urn:nbn:se:uu:diva-283080 (URN)978-91-554-9556-5 (ISBN)
Public defence
2016-06-02, B/A1:111a, BMC, Uppsala, 13:15 (English)
Available from: 2016-05-11 Created: 2016-04-11 Last updated: 2016-06-01

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Kamel, WaelOberg, DanielPunga, TanelAkusjärvi, Göran
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