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Ferrara F, Molesti E, Böttcher-Friebertshäuser E, et al. The human Transmembrane Protease Serine 2 is necessary for the production of Group 2 influenza A virus pseudotypes. J Mol Genet Med. 2013;7:309-14
Ferrara, F., Molesti, E., Böttcher-Friebertshäuser, E., Cattoli, G., Corti, D., Scott, S. D., & Temperton, N. J. (2012). The human Transmembrane Protease Serine 2 is necessary for the production of Group 2 influenza A virus pseudotypes. Journal of molecular and genetic medicine : an international journal of biomedical research, 7309-14.
Ferrara, Francesca, et al. "The human Transmembrane Protease Serine 2 is necessary for the production of Group 2 influenza A virus pseudotypes." Journal of molecular and genetic medicine : an international journal of biomedical research vol. 7 (2012): 309-14.
Ferrara F, Molesti E, Böttcher-Friebertshäuser E, Cattoli G, Corti D, Scott SD, Temperton NJ. The human Transmembrane Protease Serine 2 is necessary for the production of Group 2 influenza A virus pseudotypes. J Mol Genet Med. 2012;7:309-14. Epub 2013 Feb 20. PMID: 23577043; PMCID: PMC3614188.
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J Mol Genet Med. 2012;7:309-14. Epub 2013 Feb 20.

The human Transmembrane Protease Serine 2 is necessary for the production of Group 2 influenza A virus pseudotypes.

Journal of molecular and genetic medicine : an international journal of biomedical research

Francesca Ferrara, Eleonora Molesti, Eva Böttcher-Friebertshäuser, Giovanni Cattoli, Davide Corti, Simon D Scott, Nigel J Temperton

Affiliations

  1. Viral Pseudotype Unit, Medway School of Pharmacy, University of Kent, Central Avenue, Chatham Maritime, ME4 4TB, UK.

PMID: 23577043 PMCID: PMC3614188

Abstract

The monomer of influenza haemagglutinin is synthesized as a single polypeptide precursor that during maturation is cleaved by proteases into two active subunits. Other studies have demonstrated that the human Transmembrane Protease Serine 2 (TMPRSS2) can cleave the HA of human seasonal influenza viruses. Consequently, we have investigated the use of human Transmembrane Protease Serine 2 to produce high titre influenza haemmagglutinin (HA) lentiviral pseudotypes from Group 2 influenza viruses. Such pseudotypes represent powerful and safe tools to study viral entry and immune responses. Influenza pseudotype particles are obtained by co-transfecting human embryonic kidney HEK293T/17 cells using plasmids coding for the influenza HA, HIV gag-pol and a lentiviral vector incorporating firefly luciferase. However, in order to produce Group 2 pseudotypes, it was necessary to co-transfect a plasmid expressing the TMPRSS2 endoprotease, to achieve the necessary HA cleavage for infective particle generation. These lentiviral pseudotypes were shown to transduce HEK293T/17 cells with high efficiency. This demonstrates that TMPRSS2 is necessary for the functional activation, in vitro, of both the HA of human seasonal influenza and other Group 2 HA influenza strains. Additionally, we show that the Group 2 influenza pseudotype particles can be used as surrogate antigens in neutralization assays and are efficiently neutralized by corresponding influenza virus reference sera. These data demonstrate that the viral pseudotype system is a powerful method for serological surveillance of a wide range of influenza viruses.

Keywords: Haemagglutinin; antibody response; protease cleavage; pseudotype serology

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