Activation of the influenza virus hemagglutinin by type II transmembrane serine proteases

Abstract

The influenza virus hemagglutinin (HA) mediates viral entry into target cells. HA is synthesized as an inactive precursor and depends on proteolytic processing by host cell proteases to become active. TMPRSS2, a member of the type II transmembrane serine protease (TTSP) family, can cleave and activate HA in cell culture and was recently shown to be essential for spread and pathogenesis of influenza viruses of the H1N1 subtype in mice. In contrast, an H3N2 influenza virus was less dependent on TMPRSS2, implying that this virus may hijack other TTSPs for its activation. Moreover, studies with an animal coronavirus suggested that TMPRSS2 could impact virus infection at stages other than viral entry into cells. Therefore, the central goals of this thesis were: i) to identify TTSPs which activate HA; ii) to identify molecular determinants which control the ability of TTSPs to cleave and activate HA; iii) to determine whether TMPRSS2 promotes influenza virus infection by processes other than HA activation. The results of the present thesis show that DESC1 and MSPL, members of the TTSP family, activate diverse influenza viruses, while the related proteases TMPRSS11F, prostasin, TMPRSS11B, TMPRSS9 and TMPRSS10 fail to do so. Additionally, it was demonstrated that TTSPs which activate HA also colocalize with HA at or close to the cell membrane while this phenotype was not observed with enzymes unable to activate HA. Mutagenic analysis revealed that the stem region of TTSPs controlled their ability to colocalize and to activate HA. Moreover, evidence was obtained that targeting the scavenger receptor A domain present in the stem region of certain HA-activating TTSPs may block HA processing. Finally, evidence was obtained that TMPRSS2 and other HA-activating TTSPs can antagonize the antiviral host cell factor tetherin and might thereby promote influenza virus release form tetherin expressing cells. In sum, the present study identified two TTSPs as HA activators, and defined the stem region of TTSPs as a potential target for antiviral strategies. Finally, it was shown that TTSPs may promote influenza virus spread via two mechanisms, HA activation and tetherin inactivation.