Dear All,
I read with much interest this thoughtful thread; thanks to you all. I’ve been studying the evolution of the spike protein in detail as well.
Two issues are still open (not only) in my view: 1) the origin of nCoV-19 and 2) the origin of the furin cleavage site.
The origin of nCoV-19.
The physicist "John von Neumann used to say, with four parameters I can fit an elephant, and with five I can make him wiggle his trunk.” (quoted by Enrico Fermi, in Dayson F. Nature 2004; 427:297). Now, given that analyse Chinese railways’ timetables is far beyond my skills, I’d go instead for something more traditional: get more data. Trivial? Maybe not. I am deeply astonished by how much has been said and published on the origin(s) of nCoV-19, and all of that mainly based on the comparisons with a single sequence: Bat-CoV-RaTG13. How could this have happened?
I am used to teach students that for meaningful phylogenetic/evolutionary analyses, taxon sampling should be as balanced a possible. Here, bat relatives of the Bat-CoV-RaTG13 are clearly underrepresented. Proposed solution: Someone should ask Dr Zheng-Li Shi, where she collected that precious sample. Then send someone over there, collect some more bat samples and end once and for all this story without the need for more just-so stories. Anybody agrees with my proposal?
The origin of the furin cleavage-site.
In this thread the main proposal about the origin of the furin cleavage-site invokes replication errors and the CAGAC palindrome. Is there a chance that Gallaher’s “RNA polymerase stuttering” hypothesis could be tested in the lab? That would be terrific.
Yet, one thing is to generate (synonymous) mutations, a quite different one is to get them fixed in the population.
Yes, the S1/S2 cleavage site is present in other Coronaviruses (eg. ref to Table1-Fig2, in Coutard et al., 2020, Antiviral Res, https://doi.org/10.1016/j.antiviral.2020.104742). But not quite the same site as in 2019-nCoV, as pointed out in Walls et al. (Cell 2020, 180, 1–12). This cleavage site is “conserved among the 144 SARS CoV-2 isolates sequenced to date but not in the closely related RaTG13” (Walls 2020). The Authors demonstrate that the acquisition of the RRAR recognition site in 2019-nCoV is" something special". Indeed, the Western blot in Fig 1D shows that the S1/S2 cleavage takes place in 2019-nCoV but not in SARS-CoV: the two cleavage site are therefore not “equivalent”. Nor the cleavage takes place in a mutated sequence, in which only the Arg685 is conserved “… thereby mimicking the S1/S2 cleavage site of the related SARSr-CoV S CZX21” (Walls 2020).
Establishing the origin of the S1/S2 cleavage site in 2019-nCov is imperative for “… it is becoming increasingly apparent that proteolytic activation of spike by host cell proteases also plays a critical role … in cell and tissue tropism, host range, and pathogenesis” (Millet & Whittaker, 2015, Virus Res 202:120-134) … and “… S-protein “priming” … may provide a gain-of-function to the 2019-nCoV for efficient spreading in the human population compared to other lineage b beta-coronaviruses” (Coutard et al., 2020, Antiviral Res, https://doi.org/10.1016/j.antiviral.2020.104742).
Conclusion: To me, the origin of the furin cleavage-site in 2019-nCov is not clear yet.
Thanks again for this interesting discussion.
All the best,
Giorgio.