Transcriptional profiling reveals potential involvement of microvillous TRPM5-expressing cells in viral infection of the olfactory epithelium

  1. B. Dnate’ Baxter
  2. Eric D. Larson
  3. Laetitia Merle
  4. Paul Feinstein
  5. Arianna Gentile Polese
  6. Andrew N. Bubak
  7. Christy S. Niemeyer
  8. James Hassell
  9. Doug Shepherd
  10. Vijay R. Ramakrishnan
  11. Maria A. Nagel
  12. Diego Restrepo

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Abstract

Background

Understanding viral infection of the olfactory epithelium is essential because the olfactory nerve is an important route of entry for viruses to the central nervous system. Specialized chemosensory epithelial cells that express the transient receptor potential cation channel subfamily M member 5 (TRPM5) are found throughout the airways and intestinal epithelium and are involved in responses to viral infection.

Results

Herein we performed deep transcriptional profiling of olfactory epithelial cells sorted by flow cytometry based on the expression of mCherry as a marker for olfactory sensory neurons and for eGFP in OMP-H2B::mCherry/TRPM5-eGFP transgenic mice ( Mus musculus ). We find profuse expression of transcripts involved in inflammation, immunity and viral infection in TRPM5-expressing microvillous cells compared to olfactory sensory neurons.

Conclusion

Our study provides new insights into a potential role for TRPM5-expressing microvillous cells in viral infection of the olfactory epithelium. We find that, as found for solitary chemosensory cells (SCCs) and brush cells in the airway epithelium, and for tuft cells in the intestine, the transcriptome of TRPM5-expressing microvillous cells indicates that they are likely involved in the inflammatory response elicited by viral infection of the olfactory epithelium.

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  1. Version published to 10.1186/s12864-021-07528-y
  2. ScreenIT

    SciScore for 10.1101/2020.05.14.096016: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board StatementConsent: The TRPM5-eGFP mice and TRPM5 knockout mice(Damak et al., 2006) were obtained with written informed consent from Dr. Robert Margolskee.
    IACUC: All procedures were performed in compliance with University of Colorado Anschutz Medical Campus Institutional Animal Care and Use Committee (IACUC) that reviews the ethics of animal use.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variableBoth male and female mice were used for experiments with ages ranging from 3- 8 months.

    Table 2: Resources

    Antibodies
    SentencesResources
    At room temperature, tissue was permeabilized with Triton X-100 0.1% in PBS for 30 minutes, washed three times with PBS, blocked with Donkey serum 5% and Tween 20 0.3% in PBS for 1 hour, incubated with Chicken anti-GFP primary antibody (1:500 in blocking solution, AB_2307313 Aves labs) overnight, washed three times with PBS and incubated with Donkey anti-Chicken secondary antibody conjugated with alexa fluor 488 (1:500 in blocking solution, 703-545-155 Jackson ImmunoResearch laboratories).
    anti-GFP
    suggested: (Aves Labs Cat# GFP-1010, RRID:AB_2307313)
    anti-Chicken
    suggested: (Jackson ImmunoResearch Labs Cat# 703-545-155, RRID:AB_2340375)
    Experimental Models: Organisms/Strains
    SentencesResources
    Experiments were performed on mice from the F1 generation cross of TRPM5-eGFP and OMP-H2B::Cherry mice (OMP-H2B::mCherry/TRPM5-eGFP).
    OMP-H2B::Cherry
    suggested: None
    Olfactory epithelium cell suspensions from wild type and OMP-H2B::Cherry mice or TRPM5-eGFP mice were sorted as controls for auto fluorescence for eGFP and mCherry populations respectively.
    TRPM5-eGFP
    suggested: None
    Software and Algorithms
    SentencesResources
    RNA Sequencing Analysis: Transcript abundance was quantified from trimmed and filtered FASTQ files using Salmon v1.2.1(Patro et al., 2017) and a customized Ensembl GRCm38 (release 99) transcriptome (Zerbino et al., 2018).
    Salmon
    suggested: (Salmon, RRID:SCR_017036)
    Ensembl
    suggested: (Ensembl, RRID:SCR_002344)
    Transcript abundance was summarized at the gene level using the TxImport (Soneson et al., 2015) package in R.
    TxImport
    suggested: (tximport, RRID:SCR_016752)
    Differential gene expression was quantified using DESeq2 (Love et al., 2014) with default parameters after removing genes with an average count of < 5 reads in each group.
    DESeq2
    suggested: (DESeq, RRID:SCR_000154)
    Enrichment was calculated by dividing the number of detected genes by the number of expected genes within each ontology of the TopGO output.
    TopGO
    suggested: (topGO, RRID:SCR_014798)
    Heatmap visualization was performed using pHeatmap in R (Kolde, 2019).
    pHeatmap
    suggested: (pheatmap, RRID:SCR_016418)
    Photometrics Prime BSI) using custom Python software ((Sapoznik et al., 2020), https://github.com/qi2lab/opm).
    Python
    suggested: (IPython, RRID:SCR_001658)
    Statistical analysis: Statistical analysis was performed in Matlab (Mathworks, USA).
    Matlab
    suggested: (MATLAB, RRID:SCR_001622)

    Results from OddPub: Thank you for sharing your data.

    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: We did not find any issues relating to colormaps.

    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.

    About SciScore

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  3. Version published to 10.1101/2020.05.14.096016 on bioRxiv