The SARS-CoV-2 and other human coronavirus spike proteins are fine-tuned towards temperature and proteases of the human airways

Abstract

The high transmissibility of SARS-CoV-2 is related to abundant replication in the upper airways, which is not observed for the other highly pathogenic coronaviruses SARS-CoV and MERS-CoV. We here reveal features of the coronavirus spike (S) protein, which optimize the virus towards the human respiratory tract. First, the S proteins exhibit an intrinsic temperature preference, corresponding with the temperature of the upper or lower airways. Pseudoviruses bearing the SARS-CoV-2 spike (SARS-2-S) were more infectious when produced at 33°C instead of 37°C, a property shared with the S protein of HCoV-229E, a common cold coronavirus. In contrast, the S proteins of SARS-CoV and MERS-CoV favored 37°C, in accordance with virus preference for the lower airways. Next, SARS-2-S-driven entry was efficiently activated by not only TMPRSS2, but also the TMPRSS13 protease, thus broadening the cell tropism of SARS-CoV-2. Both proteases proved relevant in the context of authentic virus replication. TMPRSS13 appeared an effective spike activator for the virulent coronaviruses but not the low pathogenic HCoV-229E virus. Activation of SARS-2-S by these surface proteases requires processing of the S1/S2 cleavage loop, in which both the furin recognition motif and extended loop length proved critical. Conversely, entry of loop deletion mutants is significantly increased in cathepsin-rich cells. Finally, we demonstrate that the D614G mutation increases SARS-CoV-2 stability, particularly at 37°C, and, enhances its use of the cathepsin L pathway. This indicates a link between S protein stability and usage of this alternative route for virus entry. Since these spike properties may promote virus spread, they potentially explain why the spike-G614 variant has replaced the early D614 variant to become globally predominant. Collectively, our findings reveal adaptive mechanisms whereby the coronavirus spike protein is adjusted to match the temperature and protease conditions of the airways, to enhance virus transmission and pathology.

Highly Efficient SARS-CoV-2 Infection of Human Cardiomyocytes: Spike Protein-Mediated Cell Fusion and Its Inhibition

“…Notably, the ACE2 receptor was expressed at higher levels in our cardiomyocytes than in those used in the other studies. In all three studies transcripts of the proteases, cathepsins B and L, furin, and four furin-like protein convertases (PCSK5, PCSK6, PCSK7, and PCSK9) were detected, but transcripts of the proteases that enable endosome-independent viral entry in the lungs and facilitate host-to-host transmission (TMPRSS2 and TMPRSS13) ( 25 , 26 ) were below detection levels (less than 0.5 counts/million in at least 2 samples).…”

Dynamics of SARS-CoV-2 Spike Proteins in Cell Entry: Control Elements in the Amino-Terminal Domains

“…More recent studies of SARS-CoV-2 revealed cell culture adaptations that include deletions of furin protease substrates, with resultant uncleaved S proteins stabilized against S1 shedding but concomitantly less susceptible to TMPRSS2 cleavage activation of membrane fusion ( 26 , 59 , 60 ). The now-prevalent D614G change in SARS-CoV-2 also stabilizes S proteins noncovalently, promoting S1 retention ( 4 , 36 ) and extracellular infectivity ( 20 , 61 ), but potentially with reduced susceptibility to activating serine proteases ( 62 ).…”

Phosphatidylserine receptors enhance SARS-CoV-2 infection

“…ACE2-dependent coronaviruses enter cells through two different routes: 1) An endosomal route of virus uptake that requires low pH-dependent cysteine protease, cathepsin L, processing of spike and 2) a plasma membrane route that is dependent upon serine protease, TMPRSS2, cleavage of spike [3,27]. Others have reported that TMPRSS2-dependent entry is preferentially utilized by the virus when this protease is expressed [28].…”