TMPRSS2 Activates the Human Coronavirus 229E for Cathepsin-Independent Host Cell Entry and Is Expressed in Viral Target Cells in the Respiratory Epithelium

Abstract

e Infection with human coronavirus 229E (HCoV-229E) is associated with the common cold and may result in pneumonia in immunocompromised patients. The viral spike (S) protein is incorporated into the viral envelope and mediates infectious entry of HCoV-229E into host cells, a process that depends on the activation of the S-protein by host cell proteases. However, the proteases responsible for HCoV-229E activation are incompletely defined. Here we show that the type II transmembrane serine proteases TMPRSS2 and HAT cleave the HCoV-229E S-protein (229E-S) and augment 229E-S-driven cell-cell fusion, suggesting that TMPRSS2 and HAT can activate 229E-S. Indeed, engineered expression of TMPRSS2 and HAT rendered 229E-S-driven virus-cell fusion insensitive to an inhibitor of cathepsin L, a protease previously shown to facilitate HCoV-229E infection. Inhibition of endogenous cathepsin L or TMPRSS2 demonstrated that both proteases can activate 229E-S for entry into cells that are naturally susceptible to infection. In addition, evidence was obtained that activation by TMPRSS2 rescues 229E-S-dependent cell entry from inhibition by IFITM proteins. Finally, immunohistochemistry revealed that TMPRSS2 is coexpressed with CD13, the HCoV-229E receptor, in human airway epithelial (HAE) cells, and that CD13 ؉ TMPRSS2 ؉ cells are preferentially targeted by HCoV-229E, suggesting that TMPRSS2 can activate HCoV-229E in infected humans. In sum, our results indicate that HCoV-229E can employ redundant proteolytic pathways to ensure its activation in host cells. In addition, our observations and previous work suggest that diverse human respiratory viruses are activated by TMPRSS2, which may constitute a target for antiviral intervention.

Supramolecular Architecture of the Coronavirus Particle

“…One or more protease cleavage events are necessary to prime S for membrane fusion. An apparent fusion peptide of coronaviruses resides in the S2 region (Bosch et al, 2003;Sainz et al, 2005), where it is exposed by cleavage that takes place on tetraspanin-enriched membranes where host proteases including TMPRSS2 and HAT localize for some coronaviruses including MERS-CoV, while other spikes can be primed for entry by cathepsins (Bertram et al, 2013;Earnest et al, 2015;Glowacka et al, 2011;Heurich et al, 2014;Huang et al, 2006;Shulla et al, 2011;Simmons et al, 2005). This is consistent with host gene knockdown experiments that show that coronavirus entry is dependent on several elements that are important in the endosomal and lysosomal trafficking (Burkard et al, 2014;Wong et al, 2015).…”

Recent discovery and development of inhibitors targeting coronaviruses

“…Apart from this, some CoVs may also enter into the cells with the help of proteases; for example, the role of cathepsin L has been linked with the SARS-and MERS-CoVs entry, transmembrane protease serine 2 (TMPRSS2) and airway trypsin-like protease TMPRSS11D could activate the S protein for virus entry at the cell membrane during HCoV-229E and SARS-CoV infection [39][40][41].…”

“…Triazole derivatives(37)(38)(39)(40)(41)(42) against coronavirus-229E. exerted significant SARS-CoV PL pro inhibitory activity through noncompetitive inhibition.…”

The serine protease inhibitor camostat inhibits influenza virus replication and cytokine production in primary cultures of human tracheal epithelial cells

“…An indirect immunofluorescence assay was performed as reported previously [32]. Cells were fixed with 4% paraformaldehyde in PBS for 10 min at RT.…”