The type II transmembrane serine protease (TTSP) TMPRSS2 cleaves and activates the influenza virus and coronavirus surface proteins. Expression of TMPRSS2 is essential for the spread and pathogenesis of H1N1 influenza viruses in mice. In contrast, H3N2 viruses are less dependent on TMPRSS2 for viral amplification, suggesting that these viruses might employ other TTSPs for their activation. Here, we analyzed TTSPs, reported to be expressed in the respiratory system, for the ability to activate influenza viruses and coronaviruses. We found that MSPL and, to a lesser degree, DESC1 are expressed in human lung tissue and cleave and activate the spike proteins of the Middle East respiratory syndrome and severe acute respiratory syndrome coronaviruses for cell-cell and virus-cell fusion. In addition, we show that these proteases support the spread of all influenza virus subtypes previously pandemic in humans. In sum, we identified two host cell proteases that could promote the amplification of influenza viruses and emerging coronaviruses in humans and might constitute targets for antiviral intervention. IMPORTANCE Activation of influenza viruses by host cell proteases is essential for viral infectivity and the enzymes responsible are potential targets for antiviral intervention. The present study demonstrates that two cellular serine proteases, DESC1 and MSPL, activate influenza viruses and emerging coronaviruses in cell culture and, because of their expression in human lung tissue, might promote viral spread in the infected host. Antiviral strategies aiming to prevent viral activation might thus need to encompass inhibitors targeting MSPL and DESC1.

Defective interfering particles (DIPs) typically contain a highly deleted form of the viral genome, rendering them defective in virus replication. Yet upon complementation through coinfection with fully infectious standard virus (STV), interference with the viral life cycle can be observed, leading to suppressed STV replication and the release of mainly noninfectious DIPs. Interestingly, recent research indicates that DIPs may serve as an antiviral agent. Here we report the discovery of a yet-unknown type of influenza A virus-derived DIP (termed “OP7” virus) that contains numerous point mutations instead of large deletions in its genome. Furthermore, the underlying principles that render OP7 virions interfering and apparently defective seem to differ from those of conventional DIPs. In conclusion, we believe that OP7 virus might be a promising candidate for antiviral therapy. Moreover, it exerts strong effects, both on virus replication and on the host cell response, and may have been overlooked in other IAV preparations.

The influenza virus hemagglutinin (HA) facilitates viral entry into target cells. Cleavage of HA by host cell proteases is essential for viral infectivity, and the responsible enzymes are potential targets for antiviral intervention. The type II transmembrane serine protease (TTSP) TMPRSS2 has been identified as an HA activator in cell culture and in the infected host. However, it is less clear whether TMPRSS2-related enzymes can also activate HA for spread in target cells. Moreover, the activity of cellular serine protease inhibitors against HA-activating TTSPs is poorly understood. Here, we show that TMPRSS11A, another member of the TTSP family, cleaves and activates the influenza A virus (FLUAV) HA and the Middle East respiratory syndrome coronavirus spike protein (MERS-S). Moreover, we demonstrate that TMPRSS11A is expressed in murine tracheal epithelium, which is a target of FLUAV infection, and in human trachea, suggesting that the protease could support FLUAV spread in patients. Finally, we show that HA activation by the TMPRSS11A-related enzymes human airway tryptase and DESC1, but not TMPRSS11A itself, is blocked by the cellular serine protease inhibitor hepatocyte growth factor activator inhibitor type-1 (HAI-1). Our results suggest that TMPRSS11A could promote FLUAV spread in target cells and that HA-activating TTSPs exhibit differential sensitivity to blockade by cellular serine protease inhibitors.

Influenza A viruses (IAV) are commonly used to infect animal cell cultures for research purposes and vaccine production. Their replication is influenced strongly by the multiplicity of infection (MOI), which ranges over several orders of magnitude depending on the respective application. So far, mathematical models of IAV replication have paid little attention to the impact of the MOI on infection dynamics and virus yields. To address this issue, we extended an existing model of IAV replication in adherent MDCK cells with kinetics that explicitly consider the time point of cell infection. This modification does not only enable the fitting of high MOI measurements, but also the successful prediction of viral release dynamics of low MOI experiments using the same set of parameters. Furthermore, this model allows the investigation of defective interfering particle (DIP) propagation in different MOI regimes. The key difference between high and low MOI conditions is the percentage of infectious virions among the total virus particle release. Simulation studies show that DIP interference at a high MOI is determined exclusively by the DIP content of the seed virus while, in low MOI conditions, it is predominantly controlled by the de novo generation of DIPs. Overall, the extended model provides an ideal framework for the prediction and optimization of cell culture-derived IAV manufacturing and the production of DIPs for therapeutic use. Influenza is a contagious respiratory disease that severely affects several million people each year. Vaccination can provide protection against the infection, but vaccine composition has to be adjusted regularly to remain effective against this fast evolving pathogen. While influenza vaccines are mostly produced in embryonated chicken eggs, cell culture-based vaccine production is developing as an alternative providing controlled process conditions in closed systems, better scalability, and a short response time in case of pandemic outbreaks. Here, we employ a computational model to describe underlying mechanisms during the IAV infection in adherent MDCK cells. Special attention was paid on the influence of the MOI on virus spread in cell populations. Although dynamics between infections with high and low amounts of infecting virions differ significantly, our model closely captures both scenarios. Furthermore, our results provide insights into IAV-induced apoptosis and the switch from transcription to replication in intracellular IAV replication. Additionally, model simulations indicate how virus particle release is regulated, and what impact defective interfering particles have on virus replication in different infection conditions. Taken together, we developed a computational model that enables detailed analyses of IAV replication dynamics in animal cell culture.