The glycosylated extracellular domain of MUC1 protects against SARS-CoV-2 infection at the respiratory surface
Abstract
Mucins play an essential role in protecting the respiratory tract against microbial infections. The heavily O -glycosylated gel-forming mucins MUC5AC and MUC5B eliminate pathogens by mucociliary clearance while transmembrane mucins MUC1, MUC4, and MUC16 restrict microbial invasion at the apical surface of the epithelium. In this study, we determined the impact of host mucins and mucin glycans on SARS-CoV-2 spike-mediated epithelial entry. Human lung epithelial Calu-3 cells have endogenous expression of the SARS-CoV-2 entry receptor ACE2 and express high levels of glycosylated MUC1 on the surface but not MUC4 and MUC16. Removal of the MUC1 extracellular domain (ED) using the O -glycan-specific mucinase StcE greatly enhanced spike binding and viral infection. By contrast, removal of mucin glycans sialic acid and fucose did not impact viral invasion. This study implicates the glycosylated ED of MUC1 as an important component of the host defense that restricts the severity of SARS-CoV-2 infection.
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Review 3: "The glycosylated extracellular domain of MUC1 protects against SARS-CoV-2 infection at the respiratory surface"
Chatterjee et al examine the role of host mucins in SARS-CoV-2 infection and describe a role for glycosylated mucin MUC1 in restricting viral access to ACE2. The reviewers found the main claims reliable and potentially informative.
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Ieva Bagdonaite
Review 2: "The glycosylated extracellular domain of MUC1 protects against SARS-CoV-2 infection at the respiratory surface"
Chatterjee et al examine the role of host mucins in SARS-CoV-2 infection and describe a role for glycosylated mucin MUC1 in restricting viral access to ACE2. The reviewers found the main claims reliable and potentially informative.
Read the original sourceWas this evaluation helpful? -
Review 1: "The glycosylated extracellular domain of MUC1 protects against SARS-CoV-2 infection at the respiratory surface"
Chatterjee et al examine the role of host mucins in SARS-CoV-2 infection and describe a role for glycosylated mucin MUC1 in restricting viral access to ACE2. The reviewers found the main claims reliable and potentially informative.
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Strength of evidence
Reviewers: Annemieke Smet (University of Antwerp) | 📗📗📗📗◻️
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Ieva Bagdonaite (University of Copenhagen) | 📒📒📒◻️◻️
Thaher Pelaseyed (University of Gothenburg) | 📒📒📒◻️◻️Was this evaluation helpful? -
SciScore for 10.1101/2021.10.29.466408: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics not detected. Sex as a biological variable not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources For detection of the MUC1 ED, 5% mucin gels and a boric acid-Tris system were used as described previously45. α-MUC1-ED antibody 214D4 was used to detect MUC1 at a dilution of 1:1,000 in TSMT buffer. α-MUC1-EDsuggested: NoneFor detection of the CT of MUC1, 12% SDS-PAGE gel and α-MUC1-CT antibody CT2 was used. MUC1suggested: Noneα-MUC1-CTsuggested: NoneFor ACE2 detection, 10% SDS-PAGE gel and anti-ACE2 antibody (1:1,000, HPA000288, Sigma-Aldrich) was used. anti-ACE2suggested: …SciScore for 10.1101/2021.10.29.466408: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics not detected. Sex as a biological variable not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources For detection of the MUC1 ED, 5% mucin gels and a boric acid-Tris system were used as described previously45. α-MUC1-ED antibody 214D4 was used to detect MUC1 at a dilution of 1:1,000 in TSMT buffer. α-MUC1-EDsuggested: NoneFor detection of the CT of MUC1, 12% SDS-PAGE gel and α-MUC1-CT antibody CT2 was used. MUC1suggested: Noneα-MUC1-CTsuggested: NoneFor ACE2 detection, 10% SDS-PAGE gel and anti-ACE2 antibody (1:1,000, HPA000288, Sigma-Aldrich) was used. anti-ACE2suggested: (Sigma-Aldrich Cat# HPA000288, RRID:AB_1078160)Actin was detected using α-actin antibody (1:5,000; bs-0061R, Bioss). α-actinsuggested: NoneSecondary antibodies used were α-mouse IgG secondary antibody (1:10,000; A2304, Sigma), α-Armenian hamster IgG (1:10,000; GTX25745, Genetex) and α-rabbit IgG (1:10,000; A4914, Sigma). IgGsuggested: (Sigma-Aldrich Cat# A2304, RRID:AB_257993)α-Armenian hamster IgGsuggested: Noneα-rabbit IgGsuggested: NoneExperimental Models: Cell Lines Sentences Resources Cell culture: Calu-3 cells (ATCC Catalog # HTB-55), HEK-293T (ATCC Catalog # CRL-3216) and BHK-21 cells (ATCC Catalog # CCL-10) cells were routinely grown in 25 cm2 flasks in Dulbecco’s modified Eagle’s medium (DMEM) containing 10% fetal calf serum (FCS) at 37°C in 5% CO2. HEK-293Tsuggested: NoneBHK-21suggested: ATCC Cat# CCL-10, RRID:CVCL_1915)For infection experiments with the authentic SARS-CoV-2 virus, Calu-3 cells were prepared as described above and inoculated with approximately 200 pfu of SARS-CoV-2. Calu-3suggested: NoneSoftware and Algorithms Sentences Resources Dimensionality reduction was done using the Seurat Package42 in Rstudio (version 1.2.5019), starting with a principle component analysis. Rstudiosuggested: (RStudio, RRID:SCR_000432)Immunofluorescent stainings were performed as described for SARS-CoV-2 stock production and scanned plates were analyzed using ImageQuant TL software. ImageQuantsuggested: (ImageQuant, RRID:SCR_014246)The GraphPad Prism 9 software package was used for all statistical analyses. GraphPadsuggested: (GraphPad Prism, RRID:SCR_002798)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).
Results from JetFighter: Please consider improving the rainbow (“jet”) colormap(s) used on pages 17, 19 and 20. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
Results from scite Reference Check: We found no unreliable references.
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