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01.12.2018 | Research | Ausgabe 1/2018 Open Access

Virology Journal 1/2018

Bacterial ribonuclease binase exerts an intra-cellular anti-viral mode of action targeting viral RNAs in influenza a virus-infected MDCK-II cells

Zeitschrift:
Virology Journal > Ausgabe 1/2018
Autoren:
Raihan Shah Mahmud, Ahmed Mostafa, Christin Müller, Pumaree Kanrai, Vera Ulyanova, Yulia Sokurenko, Julia Dzieciolowski, Irina Kuznetsova, Olga Ilinskaya, Stephan Pleschka
Wichtige Hinweise

Electronic supplementary material

The online version of this article (https://​doi.​org/​10.​1186/​s12985-017-0915-1) contains supplementary material, which is available to authorized users.

Abstract

Background

Influenza is a severe contagious disease especially in children, elderly and immunocompromised patients. Beside vaccination, the discovery of new anti-viral agents represents an important strategy to encounter seasonal and pandemic influenza A virus (IAV) strains. The bacterial extra-cellular ribonuclease binase is a well-studied RNase from Bacillus pumilus. Treatment with binase was shown to improve survival of laboratory animals infected with different RNA viruses. Although binase reduced IAV titer in vitro and in vivo, the mode of action (MOA) of binase against IAV at the molecular level has yet not been studied in depth and remains elusive.

Methods

To analyze whether binase impairs virus replication by direct interaction with the viral particle we applied a hemagglutination inhibition assay and monitored the integrity of the viral RNA within the virus particle by RT-PCR. Furthermore, we used Western blot and confocal microscopy analysis to study whether binase can internalize into MDCK-II cells. By primer extension we examined the effect of binase on the integrity of viral RNAs within the cells and using a mini-genome system we explored the effect of binase on the viral expression.

Results

We show that (i) binase does not to attack IAV particle-protected viral RNA, (ii) internalized binase could be detected within the cytosol of MDCK-II cells and that (iii) binase impairs IAV replication by specifically degrading viral RNA species within the infected MDCK-II cells without obvious effect on cellular mRNAs.

Conclusion

Our data provide novel evidence suggesting that binase is a potential anti-viral agent with specific intra-cellular MOA.
Zusatzmaterial
Additional file 1: Figure S1. The effect of binase on HA inhibition (HI). Twofold serially diluted binase (105 U/ml) was incubated with 4HAU of H1N1pdm09 and then supplemented with 1% chicken erythrocytes (RBCs) to detect the binase/ligand interaction (binase + H1N1pdm + RBCs). Meanwhile, twofold serially diluted binase (105 U/ml) was incubated with 1% chicken erythrocytes (RBCs) and then 4 HAU of H1N1pdm09 were added to explore the binase/receptor interaction (binase + RBCs + H1N1pdm). HI assay was conducted in triplicate for each serial dilution (1:10–1:1280).1xPBS, binase (105 U/ml), and H1N1pdm09 (4 HAU) were used as controls. (PDF 432 kb)
12985_2017_915_MOESM1_ESM.pdf
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