Dataset Information

Expansion of Single Cell Transcriptomics Data of SARS-CoV Infection in Human Bronchial Epithelial Cells to COVID-19.

ABSTRACT:

Background

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 19 (COVID-19) that was emerged as a new member of coronaviruses since December 2019 in Wuhan, China and then after was spread in all continentals. Since SARS-CoV-2 has shown about 77.5% similarity to SARS-CoV, the transcriptome and immunological regulations of SARS-CoV-2 was expected to have high percentage of overlap with SARS-CoV.

Results

In this study, we applied the single cell transcriptomics data of human bronchial epithelial cells (2B4 cell line) infected with SARS-CoV, which was annotated in the Expression Atlas database to expand this data to COVID-19. In addition, we employed system biology methods including gene ontology (GO) and Reactome pathway analyses to define functional genes and pathways in the infected cells with SARS-CoV. The transcriptomics analysis on the Expression Atlas database revealed that most genes from infected 2B4 cell line with SARS-CoV were downregulated leading to immune system hyperactivation, induction of signaling pathways, and consequently a cytokine storm. In addition, GO:0016192 (vesicle-mediated transport), GO:0006886 (intracellular protein transport), and GO:0006888 (ER to Golgi vesicle-mediated transport) were shown as top three GOs in the ontology network of infected cells with SARS-CoV. Meanwhile, R-HAS-6807070 (phosphatase and tensin homolog or PTEN regulation) showed the highest association with other Reactome pathways in the network of infected cells with SARS-CoV. PTEN plays a critical role in the activation of dendritic cells, B- and T-cells, and secretion of proinflammatory cytokines, which cooperates with downregulated genes in the promotion of cytokine storm in the COVID-19 patients.

Conclusions

Based on the high similarity percentage of the transcriptome of SARS-CoV with SARS-CoV-2, the data of immunological regulations, signaling pathways, and proinflammatory cytokines in SARS-CoV infection can be expanded to COVID-19 to have a valid platform for future pharmaceutical and vaccine studies.

SUBMITTER: Zolfaghari Emameh R

PROVIDER: S-EPMC7377208 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Severe acute respiratory syndrome-associated coronavirus (SARS-CoV) infection causes an immune-mediated disease. We have recently shown that SARS-CoV-induced epithelial Calu-3 cytokines could exacerbate and dampen host inflammatory and T cell responses, respectively, through modulating the functions of macrophages and dendritic cells, thereby suggesting that not only are lung epithelial cells the primary cells of SARS-CoV infection, but they also involve in initiating and orchestrating the host innate and adaptive immunity. Comprehensive evaluation of the complex epithelial signaling to SARS-CoV is, thus, crucial for paving the way to better understand SARS pathogenesis and develop the innovative therapeutics against SARS. Here, based on the microarray-based functional genomics, we reported that 2B4 cells, a clonal derivative of Calu-3 cells, elicited a temporal and spatial activation of nuclear factor (NF)kappaB, activator protein (AP)-1 (ATF2/c-Jun), and interferon regulatory factor (IRF)-3/-7 at 12-, 24-, and 48-hrs post infection (p.i.), respectively, resulting in the activation of many antiviral genes, including interferon (IFN)-Î², -Î»s, SARS-related inflammatory mediators, and various IFN-stimulated genes (ISGs). While elevated responses of IFN-Î² and IFN-Î»s were not detected until 48-hrs p.i., as a consequence of a delayed IRF-3/-7 activation, we showed, for the first time, that both types of IFNs exerted previously under-described non-redundant, complementary, and/or synergistic effects on the epithelial defense against SARS-CoV. Collectively, our results highlight the molecular mechanisms of the sequential activation of virus- and IFN-dependent signaling of lung epithelial cells against SARS-CoV and identify novel cellular targets for future studies, aiming at advancing strategies against SARS. DNA microarrays were performed in the Molecular Genomics (MG) Core Facilities at UTMB. The detailed information about the procedures of microarray analysis has been posted on the MG Core Facilities website (genomics@scms.utmb.edu). To characterize the dynamic, spatial, and temporal changes of the gene expression induced by SARS-CoV, confluent 2B4 cells grown in T-75 flasks were infected with SARS-CoV (MOI=0.1) or remained uninfected (as control) for 12, 24, and 48hrs. Because 2B4 cells were also permissive to the productive infection of Dhori virus (DHOV), a member of the Orthomyxoviridae family within the Thogotovirus genus, resulting in robust responses of IFNs and other pro-inflammatory mediators, we also established parallel cultures of DHOV-infected 2B4 cells (MOI=0.1) for the comparative analysis of global gene expression elicited by SARS-CoV- versus DHOV-infected 2B4 cells. To meet the minimal number required for application of statistical algorithms, we performed the study in triplicate at each time point for mock-, SARS-CoV-, and DHOV-infected cultures, yielding a total of 27 arrays. Briefly, supernatants were harvested from differentially treated cultures at 12-, 24-, and 48-hrs p.i. for subsequent analyses of viral yields and cytokine profiling, whereas the cells were subjected to total RNA extraction by using an RNAqueous-4PCR kit and following the protocol recommended by the manufacturer (Ambion, Austin, TX). Purified RNA samples were sent to our core facility for conversion to cDNA, biotin-labeled, and hybridized to 27 Affymetrix Human Genome U133 Plus 2.0 âGene Chipsâ each of which contained 54,675 probe set identifiers representing more than ~47,400 transcripts that identify ~38,500 well-characterized genes, and various internal controls (Affymetrix, Santa Clara, CA). Mock-infected cells were compared to cells infected with SARS-CoV or DHOV at each time point.

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