Skip to main content

Haemagglutinin activation by human transmembrane protease serine 2 or by human airway trypsin-like protease is necessary for the production of high titre influenza A virus pseudotypes that can be employed for the evaluation of pandemic potential

Ferrara, Francesca, Molesti, Eleonora, Böttcher-Friebertshäuser, Eva, Corti, Davide, Cattoli, Giovanni, Scott, Simon D., Temperton, Nigel J. (2013) Haemagglutinin activation by human transmembrane protease serine 2 or by human airway trypsin-like protease is necessary for the production of high titre influenza A virus pseudotypes that can be employed for the evaluation of pandemic potential. In: Options for the Control of Influenza VIII, 5-10 Sep 2013, Cape Town. (doi:P2-493) (KAR id:35142)

Abstract

Background: The monomer of influenza haemagglutinin (HA) is synthesized as a single polypeptide precursor that during maturation is cleaved by proteases into two active subunits. Recent studies have demonstrated that the human transmembrane protease serine 2 (TMPRSS2) and the human airway trypsin-like protease (HAT) can activate, by cleavage, the HA of human seasonal influenza viruses. As a model of activation of all influenza subtypes (human H1, H2, H3 viruses and other representative viruses from group 1 and group 2 avian subtypes), we have investigated the use of human TMPRSS2 and HAT to produce high titre influenza HA lentiviral pseudotypes. Such pseudotypes represent powerful and safe tools to study viral entry mechanisms and zoonotic potential. Materials and Methods: Influenza pseudotype particles are obtained by cotransfecting human embryonic kidney HEK293T/17 cells using plasmids coding for the influenza HA, HIV gag-pol and a retroviral vector incorporating firefly luciferase. To investigate the role of these proteases in the HA activation necessary for infective particle generation, a plasmid expressing TMPRSS2 or HAT was cotransfected during pseudotype production. The HA lentiviral pseudotypes produced were used to investigate cell entry potential via transduction of target cell lines and as surrogate antigens in neutralization assays. Results: Influenza pseudotype particles produced by cotransfection of these proteases can transduce HEK293T/17 cells with high efficiency compared with the pseudotypes produced in the absence of proteases. The high titre of these influenza pseudotypes permits their use as surrogate antigens in neutralization assays and subtype-specific sera can readily neutralize them. Conclusions: TMPRSS2 and/or HAT can activate, in vitro, both the HA of human seasonal influenza and also other avian HA influenza strains in a pseudotype particle production system. Furthermore, this panel of influenza pseudotypes can be used in neutralization assays to study heterosubtypic antibody responses and pandemic potential.

Item Type: Conference or workshop item (Poster)
DOI/Identification number: P2-493
Subjects: Q Science > QR Microbiology > QR355 Virology
Divisions: Divisions > Division of Natural Sciences > Medway School of Pharmacy
Depositing User: Nigel Temperton
Date Deposited: 07 Sep 2013 11:20 UTC
Last Modified: 16 Nov 2021 10:12 UTC
Resource URI: https://kar.kent.ac.uk/id/eprint/35142 (The current URI for this page, for reference purposes)

University of Kent Author Information

Ferrara, Francesca.

Creator's ORCID:
CReDIT Contributor Roles:

Molesti, Eleonora.

Creator's ORCID:
CReDIT Contributor Roles:

Scott, Simon D..

Creator's ORCID: https://orcid.org/0000-0002-8290-0461
CReDIT Contributor Roles:

Temperton, Nigel J..

Creator's ORCID: https://orcid.org/0000-0002-7978-3815
CReDIT Contributor Roles:
  • Depositors only (login required):

Total unique views for this document in KAR since July 2020. For more details click on the image.