Gene Expression
Nebulas
Gene Expression NebulasSummary: Pre-existing memory T-cells against SARS-CoV-2 are present in a fraction of unexposed individuals and their induction by common cold corona viruses (CCCoVs) infection is suggested. Here we demonstrate that SARS-CoV-2-reactive T-cells were present in the memory compartment of virtually all unexposed individuals but possessed only low functional avidity. They harbored multiple, highly variable cross-reactivities that were not restricted to CCCoVs. Cross-reactivity to CCCoV was almost absent in COVID-19 patients. This was irrespective of strong T-cell memory against CCCoV in all donors. In severe but not mild COVID-19, SARS-CoV-2-specific T-cells also displayed low functional avidity and reduced clonal expansion, despite strongly increased frequencies. Our data question a major protective role of CCCoV for COVID-19. Instead, we suggest that a low avidity pre-existing T-cell memory may contribute to the excessive but low avidity T-cell responses, which we identified here as a hallmark of severe COVID-19
Overall Design: Single-cell sequencing of 6 unexposed and 14 COVID-19 patients
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| Treatment Protocol: | PBMCs were stimulated for 7h with peptide pools of SARS-CoV-2 and reactive CD154+CD4+ T cells were isolated |
| Extract Protocol: | CD154+ cells were isolated by MACS and further purified by FACS sorting on a MACSQuant Tyto (Miltenyi Biotec) based on dual expression of CD154 and CD69. Sorted CD154+ T cells were removed from the sorting chamber into pre-coated low-bind collection tubes, 1ml RPMI-1640 medium supplemented with 5% AB Serum was added, and cells were centrifuged for 5 min at 400 x g, 4°C. The supernatant was carefully removed leaving 10-30μl to reach a maximum concentration of 1000 cells /μl. |
| Library Construction Protocol: | Single-cell suspensions were loaded on a ChromiumChip G (10x Genomics) according to the manufacturer’s instructions for processing with the Chromium Next GEM Single Cell 5′ Library and Gel Bead Kit v1.1. TCR single-cell libraries were subsequently prepared from the same cells with the Chromium Single Cell V(D)J Enrichment Kit, Human T Cell. Chromium Next GEM Single Cell 5′ Library |
| Molecule Type: | Poly(A)+ RNA |
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| Library Layout: | PAIRED |
| Library Strand: | - |
| Platform: | ILLUMINA |
| Instrument Model: | Illumina NovaSeq 6000 |
| Strand-Specific: | - |
| Data Resource | GEN Sample ID | GEN Dataset ID | Project ID | BioProject ID | Sample ID | Sample Name | BioSample ID | Sample Accession | Experiment Accession | Release Date | Submission Date | Update Date | Species | Race | Ethnicity | Age | Age Unit | Gender | Source Name | Tissue | Cell Type | Cell Subtype | Cell Line | Disease | Disease State | Development Stage | Mutation | Phenotype | Case Detail | Control Detail | Growth Protocol | Treatment Protocol | Extract Protocol | Library Construction Protocol | Molecule Type | Library Layout | Strand-Specific | Library Strand | Spike-In | Strategy | Platform | Instrument Model | Cell Number | Reads Number | Gbases | AvgSpotLen1 | AvgSpotLen2 | Uniq Mapping Rate | Multiple Mapping Rate | Coverage Rate |
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