The Sars-Cov-2 And Other Human Coronavirus Spike Proteins Are Fine-Tuned Towards Temperature And Proteases Of The Human Airways

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 degrees C instead of 37 degrees 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 degrees 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 degrees 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.Author summaryThe devastating COVID-19 pandemic is caused by SARS-CoV-2, a novel virus that despite recent zoonotic introduction is already very well adapted to its human host. Its rapid spread is related to abundant replication in the upper airways, which is not observed for other highly pathogenic human coronaviruses. To understand the role of the viral spike protein in this airway adaptation, we constructed pseudoviruses of SARS-CoV-2 and other coronaviruses that cause severe pneumonia or, on the contrary, a mild common cold. The key findings were verified with authentic virus. We reveal features of the spike proteins, which optimize the coronavirus towards specific parts of the respiratory tract. Namely, we show that the spike proteins exhibit intrinsic temperature preference to precisely match the upper (similar to 33 degrees C) or lower (37 degrees C) airways. We recognized which proteases of human airways activate the spike for virus entry, in particular one protease that may mediate coronavirus virulence. Finally, a link was perceived between spike stability and entry via endosomal proteases. We propose that these mechanisms of spike fine-tuning may have contributed to a global shift in SARS-CoV-2 epidemiology, from the early spike-D614 to the currently predominating G614 variant.