MOLECULAR DETERMINANTS of SARS-CoV-2 CELLULAR TROPISM in VITRO
Topics in Antiviral Medicine
; 30(1 SUPPL):62, 2022.
Article
in English
| EMBASE | ID: covidwho-1880746
ABSTRACT
Background:
Lung cell lines to model SARS-CoV-2 replication in vitro are greatly limited hampering the rigorous study of SARS-CoV-2-host interactions. We analyzed a panel of 10 airway cell lines with various levels of ACE2 expression to identify models of SARS-CoV-2 infection. We found that none of the ACE2 expressing cell lines supported replication, whereas the H522 human lung adenocarcinoma cells were naturally permissive to SARS-CoV-2 infection despite detectable expression of ACE2. We confirmed that SARS-CoV-2 replication is indeed completely independent of ACE2 in H522s but dependent on heparan sulfates and the E484D substitution within the Spike. Further, we show that many of the ACE2 positive non-permissive cell lines express high basal levels of interferon-stimulated genes, which can be overcome by inhibition of the JAK/STAT pathway or by ACE2 overexpression. Together, our findings highlight ACE2-independent pathways can control the cellular tropism of SARS-CoV-2.Methods:
Conventional molecular virology assays have been conducted to study the permissiveness of a panel of 10 cell lines expressing various levels of ACE2. ACE2 independence of SARS-CoV-2 replication was validated by antibody blocking, Fc-ACE2 decoy peptide and CRISPR-based approaches in H522 cells. RNA sequencing was used to study the basal level of genes in the type-I IFN pathway in the panel of 10 cell lines, which was further validated by western blotting and qRT-PCR. A panel of 5 cell lines, with varying expression levels of ACE2 and TMPRSS2, were pre-treated with Ruxolitinib, a JAK inhibitor, and infected with SARS-CoV-2 strain 2019-nCoV/USA-WA1/2020 and spike variants. Viral replication was detected through analysis of cell associated RNAResults:
H522 human lung adenocarcinoma supports SARS-CoV-2 replication in a completely ACE2-independent manner. Transcriptomic analysis revealed basal high level of expression of interferon response pathway genes in some ACE2-positive cells recalcitrant to SARS-CoV-2 infection. Infection of OE21 and SCC25 cells required blocking of the IFN response pathway or ACE2 overexpression to allow SARS-CoV-2 infection.Conclusion:
These findings suggest that SARS-CoV-2 replication can proceed in complete absence of ACE2 and that the innate immunity is a key determinant of SARS-CoV-2 cellular tropism. These findings may explain the complex SARS-CoV-2 pathogenesis in vivo as it shows that factors independent of ACE2 can define cellular tropism.
endogenous compound; interferon; Janus kinase; Janus kinase inhibitor; ruxolitinib; STAT protein; transmembrane protease serine 2; airway cell; animal cell; animal experiment; animal model; cancer model; clustered regularly interspaced short palindromic repeat; conference abstract; controlled study; coronavirus disease 2019; gene expression; gene overexpression; human; in vitro study; in vivo study; innate immunity; lung adenocarcinoma; NCI-H522 cell line; nonhuman; polymerase chain reaction; protein expression; RNA sequencing; SARS-CoV-2 (clinical isolate USA/WA1/2020); Severe acute respiratory syndrome coronavirus 2; signal transduction; spike; tropism; virology; virus replication; Western blotting
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Collection:
Databases of international organizations
Database:
EMBASE
Language:
English
Journal:
Topics in Antiviral Medicine
Year:
2022
Document Type:
Article
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