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Published: November 12, 2023
DOI:https://doi.org/10.1016/j.ebiom.2023.104869
Bing Han,a,f Yibing Lv,b,f Dominique Moser,a,f Xiaoqi Zhou,b Tobias Woehrle,a Lianyong Han,c Andreas Osterman,d Martina Rudelius,eAlexanderChoukér,a,∗,f andPingLeib,∗∗,f
Summary
Background
SARS-CoV-2 infects host cells via an ACE2/TMPRSS2 entry mechanism. Monocytes and macrophages, which play a key role during severe COVID-19 express only low or no ACE2, suggesting alternative entry mechanisms in these cells. In silico analyses predicted GRP78, which is constitutively expressed on monocytes and macrophages, to be a potential candidate receptor for SARS-CoV-2 virus entry.
Methods
Hospitalized COVID-19 patients were characterized regarding their pro-inflammatory state and cell surface GRP78 (csGRP78) expression in comparison to healthy controls. RNA from CD14+monocytes of patients and controls were subjected to transcriptome analysis that was specifically complemented by bioinformatic re-analyses of bronchoalveolar lavage fluid (BALF) datasets of COVID-19 patients with a focus on monocyte/macrophage subsets, SARS-CoV-2 infection state as well as GRP78 gene expression. Monocyte and macrophage immunohistocytochemistry on GRP78 was conducted in post-mortem lung tissues. SARS-CoV-2 spike and GRP78 protein interaction was analyzed by surface plasmon resonance, GST Pull-down and Co-Immunoprecipitation. SARS-CoV-2 pseudovirus or single spike protein uptake was quantified in csGRP78high THP-1 cells.
Findings
Cytokine patterns, monocyte activation markers and transcriptomic changes indicated typical COVID-19 associated inflammation accompanied by upregulated csGRP78 expression on peripheral blood and lung monocytes/macrophages. Subsequent cell culture experiments confirmed an association between elevated pro-inflammatory cytokine levels and upregulation of csGRP78. Interaction of csGRP78 and SARS-CoV-2 spike protein with a dissociation constant of KD = 55.2 nM was validated in vitro. Infection rate analyses in ACE2low and GRP78high THP-1 cells showed increased uptake of pseudovirus expressing SARS-CoV-2 spike protein.
Interpretation
Our results demonstrate that csGRP78 acts as a receptor for SARS-CoV-2 spike protein to mediate ACE2-independent virus entry into monocytes.
DOI:https://doi.org/10.1016/j.ebiom.2023.104869
Bing Han,a,f Yibing Lv,b,f Dominique Moser,a,f Xiaoqi Zhou,b Tobias Woehrle,a Lianyong Han,c Andreas Osterman,d Martina Rudelius,eAlexanderChoukér,a,∗,f andPingLeib,∗∗,f
Summary
Background
SARS-CoV-2 infects host cells via an ACE2/TMPRSS2 entry mechanism. Monocytes and macrophages, which play a key role during severe COVID-19 express only low or no ACE2, suggesting alternative entry mechanisms in these cells. In silico analyses predicted GRP78, which is constitutively expressed on monocytes and macrophages, to be a potential candidate receptor for SARS-CoV-2 virus entry.
Methods
Hospitalized COVID-19 patients were characterized regarding their pro-inflammatory state and cell surface GRP78 (csGRP78) expression in comparison to healthy controls. RNA from CD14+monocytes of patients and controls were subjected to transcriptome analysis that was specifically complemented by bioinformatic re-analyses of bronchoalveolar lavage fluid (BALF) datasets of COVID-19 patients with a focus on monocyte/macrophage subsets, SARS-CoV-2 infection state as well as GRP78 gene expression. Monocyte and macrophage immunohistocytochemistry on GRP78 was conducted in post-mortem lung tissues. SARS-CoV-2 spike and GRP78 protein interaction was analyzed by surface plasmon resonance, GST Pull-down and Co-Immunoprecipitation. SARS-CoV-2 pseudovirus or single spike protein uptake was quantified in csGRP78high THP-1 cells.
Findings
Cytokine patterns, monocyte activation markers and transcriptomic changes indicated typical COVID-19 associated inflammation accompanied by upregulated csGRP78 expression on peripheral blood and lung monocytes/macrophages. Subsequent cell culture experiments confirmed an association between elevated pro-inflammatory cytokine levels and upregulation of csGRP78. Interaction of csGRP78 and SARS-CoV-2 spike protein with a dissociation constant of KD = 55.2 nM was validated in vitro. Infection rate analyses in ACE2low and GRP78high THP-1 cells showed increased uptake of pseudovirus expressing SARS-CoV-2 spike protein.
Interpretation
Our results demonstrate that csGRP78 acts as a receptor for SARS-CoV-2 spike protein to mediate ACE2-independent virus entry into monocytes.
