The Functional Consequences of the Novel Ribosomal Pausing Site in SARS-CoV-2 Spike Glycoprotein RNA
The SARS-CoV-2 Spike glycoprotein (S protein) acquired a unique new 4 amino acid -PRRA- insertion sequence at amino acid residues (aa) 681–684 that forms a new furin cleavage site in S protein as well as several new glycosylation sites. We studied various statistical properties of the -PRRA- inserti...
Main Authors: | , , , , , , , |
---|---|
Format: | Article |
Language: | English |
Published: |
MDPI AG
2021-06-01
|
Series: | International Journal of Molecular Sciences |
Subjects: | |
Online Access: | https://www.mdpi.com/1422-0067/22/12/6490 |
id |
doaj-0a8bc8b857ed4f0887dec003129626ac |
---|---|
record_format |
Article |
spelling |
doaj-0a8bc8b857ed4f0887dec003129626ac2021-07-01T00:24:43ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672021-06-01226490649010.3390/ijms22126490The Functional Consequences of the Novel Ribosomal Pausing Site in SARS-CoV-2 Spike Glycoprotein RNAOlga A. Postnikova0Sheetal Uppal1Weiliang Huang2Maureen A. Kane3Rafael Villasmil4Igor B. Rogozin5Eugenia Poliakov6T. Michael Redmond7Laboratory of Retinal Cell & Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USALaboratory of Retinal Cell & Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USADepartment of Pharmaceutical Sciences, School of Pharmacy Mass Spectrometry Center, University of Maryland, Baltimore, MD 21201, USADepartment of Pharmaceutical Sciences, School of Pharmacy Mass Spectrometry Center, University of Maryland, Baltimore, MD 21201, USAFlow Cytometry Core Facility, National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USANational Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD 20894, USALaboratory of Retinal Cell & Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USALaboratory of Retinal Cell & Molecular Biology, National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USAThe SARS-CoV-2 Spike glycoprotein (S protein) acquired a unique new 4 amino acid -PRRA- insertion sequence at amino acid residues (aa) 681–684 that forms a new furin cleavage site in S protein as well as several new glycosylation sites. We studied various statistical properties of the -PRRA- insertion at the RNA level (CCUCGGCGGGCA). The nucleotide composition and codon usage of this sequence are different from the rest of the SARS-CoV-2 genome. One of such features is two tandem CGG codons, although the CGG codon is the rarest codon in the SARS-CoV-2 genome. This suggests that the insertion sequence could cause ribosome pausing as the result of these rare codons. Due to population variants, the Nextstrain divergence measure of the CCU codon is extremely large. We cannot exclude that this divergence might affect host immune responses/effectiveness of SARS-CoV-2 vaccines, possibilities awaiting further investigation. Our experimental studies show that the expression level of original RNA sequence “wildtype” spike protein is much lower than for codon-optimized spike protein in all studied cell lines. Interestingly, the original spike sequence produces a higher titer of pseudoviral particles and a higher level of infection. Further mutagenesis experiments suggest that this dual-effect insert, comprised of a combination of overlapping translation pausing and furin sites, has allowed SARS-CoV-2 to infect its new host (human) more readily. This underlines the importance of ribosome pausing to allow efficient regulation of protein expression and also of cotranslational subdomain folding.https://www.mdpi.com/1422-0067/22/12/6490ribosome stallingSARS-CoV-2spike proteincodon usageribosome pausing site |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Olga A. Postnikova Sheetal Uppal Weiliang Huang Maureen A. Kane Rafael Villasmil Igor B. Rogozin Eugenia Poliakov T. Michael Redmond |
spellingShingle |
Olga A. Postnikova Sheetal Uppal Weiliang Huang Maureen A. Kane Rafael Villasmil Igor B. Rogozin Eugenia Poliakov T. Michael Redmond The Functional Consequences of the Novel Ribosomal Pausing Site in SARS-CoV-2 Spike Glycoprotein RNA International Journal of Molecular Sciences ribosome stalling SARS-CoV-2 spike protein codon usage ribosome pausing site |
author_facet |
Olga A. Postnikova Sheetal Uppal Weiliang Huang Maureen A. Kane Rafael Villasmil Igor B. Rogozin Eugenia Poliakov T. Michael Redmond |
author_sort |
Olga A. Postnikova |
title |
The Functional Consequences of the Novel Ribosomal Pausing Site in SARS-CoV-2 Spike Glycoprotein RNA |
title_short |
The Functional Consequences of the Novel Ribosomal Pausing Site in SARS-CoV-2 Spike Glycoprotein RNA |
title_full |
The Functional Consequences of the Novel Ribosomal Pausing Site in SARS-CoV-2 Spike Glycoprotein RNA |
title_fullStr |
The Functional Consequences of the Novel Ribosomal Pausing Site in SARS-CoV-2 Spike Glycoprotein RNA |
title_full_unstemmed |
The Functional Consequences of the Novel Ribosomal Pausing Site in SARS-CoV-2 Spike Glycoprotein RNA |
title_sort |
functional consequences of the novel ribosomal pausing site in sars-cov-2 spike glycoprotein rna |
publisher |
MDPI AG |
series |
International Journal of Molecular Sciences |
issn |
1661-6596 1422-0067 |
publishDate |
2021-06-01 |
description |
The SARS-CoV-2 Spike glycoprotein (S protein) acquired a unique new 4 amino acid -PRRA- insertion sequence at amino acid residues (aa) 681–684 that forms a new furin cleavage site in S protein as well as several new glycosylation sites. We studied various statistical properties of the -PRRA- insertion at the RNA level (CCUCGGCGGGCA). The nucleotide composition and codon usage of this sequence are different from the rest of the SARS-CoV-2 genome. One of such features is two tandem CGG codons, although the CGG codon is the rarest codon in the SARS-CoV-2 genome. This suggests that the insertion sequence could cause ribosome pausing as the result of these rare codons. Due to population variants, the Nextstrain divergence measure of the CCU codon is extremely large. We cannot exclude that this divergence might affect host immune responses/effectiveness of SARS-CoV-2 vaccines, possibilities awaiting further investigation. Our experimental studies show that the expression level of original RNA sequence “wildtype” spike protein is much lower than for codon-optimized spike protein in all studied cell lines. Interestingly, the original spike sequence produces a higher titer of pseudoviral particles and a higher level of infection. Further mutagenesis experiments suggest that this dual-effect insert, comprised of a combination of overlapping translation pausing and furin sites, has allowed SARS-CoV-2 to infect its new host (human) more readily. This underlines the importance of ribosome pausing to allow efficient regulation of protein expression and also of cotranslational subdomain folding. |
topic |
ribosome stalling SARS-CoV-2 spike protein codon usage ribosome pausing site |
url |
https://www.mdpi.com/1422-0067/22/12/6490 |
work_keys_str_mv |
AT olgaapostnikova thefunctionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT sheetaluppal thefunctionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT weilianghuang thefunctionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT maureenakane thefunctionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT rafaelvillasmil thefunctionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT igorbrogozin thefunctionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT eugeniapoliakov thefunctionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT tmichaelredmond thefunctionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT olgaapostnikova functionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT sheetaluppal functionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT weilianghuang functionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT maureenakane functionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT rafaelvillasmil functionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT igorbrogozin functionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT eugeniapoliakov functionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna AT tmichaelredmond functionalconsequencesofthenovelribosomalpausingsiteinsarscov2spikeglycoproteinrna |
_version_ |
1721348696692490240 |