SARS-CoV-2 mutations affect proteasome processing to alter CD8+ T cell responses (preprint)

Viral CD8+ epitopes are generated by the cellular turnover of viral proteins, predominantly by the proteasome. Mutations located within viral epitopes can result in escape from memory T cells but the contribution of mutations in flanking regions of epitopes in SARS-CoV-2 has not been investigated. Focusing on two of the most dominant SARS-CoV-2 nucleoprotein CD8+ epitopes, we identified mutations in epitope flanking regions and investigated the contribution of these mutations to antigen processing and T cell activation using SARS-CoV-2 nucleoprotein transduced B cell lines and in vitro proteasomal processing of peptides. We found that decreased NP9-17-B*27:05 CD8+ T cell responses to the NP-Q7K mutation correlated with lower epitope surface expression, likely due to a lack of efficient epitope production by the proteasome, suggesting immune escape caused by this mutation. In contrast, NP-P6L and NP-D103N/Y mutations flanking the NP9-17-B*27:05 and NP105-113-B*07:02 epitopes, respectively, increased CD8+ T cell responses associated with enhanced epitope production by the proteasome. Our results provide evidence that SARS-CoV-2 mutations outside the epitope could have a significant impact on antigen processing and presentation, thereby contributing to escape from immunodominant T cell responses. Alternatively, mutations could enhance antigen processing and efficacy of T cell recognition, opening new avenues for improving future vaccine designs.

Memory cytotoxic SARS-CoV-2 spike protein-specific CD4+ T cells associate with viral control (preprint)

Little is known of the role of cytotoxic CD4+ T-cells in the control of viral replication. Here, we investigate CD4+ T-cell responses to three dominant SARS-CoV-2 epitopes and evaluate antiviral activity, including cytotoxicity and antiviral cytokine production. Diverse T cell receptor (TCR) usage including public TCRs were identified; surprisingly, cytotoxic CD4+ T-cells were found to have signalling and cytotoxic pathways distinct from classical CD8+ T-cells, with increased expression of chemokines and tissue homing receptors promoting migration. We show the presence of cytolytic CD4+ T-cells during primary infection associates with COVID-19 disease severity. Robust immune memory 6-9 months post-infection or vaccination provides CD4+ T-cells with potent antiviral activity. Our data support a model where CD4+ killer cells drive immunopathogenesis during primary infection and CD4+ memory responses are protective during secondary infection. Our study highlights the unique features of cytotoxic CD4+ T-cells that use distinct functional pathways, providing preventative and therapeutic opportunities.

Immunodominant NP105-113-B*0702 cytotoxic T cell response controls viral replication and is associated with less severe COVID-19 disease (preprint)

NP 105-113 -B*07:02 specific CD8 + T-cell responses are considered among the most dominant in SARS-CoV-2-infected individuals. We found strong association of this response with mild disease. Analysis of NP 105-113 -B*07:02 specific T-cell clones and single cell sequencing were performed concurrently, with functional avidity and anti-viral efficacy assessed using an in vitro SARS-CoV-2 infection system, and were correlated with TCR usage, transcriptome signature, and disease severity (acute N=77, convalescent N=52). We demonstrated a beneficial association of NP 105-113 -B*07:02 specific T-cells in COVID-19 disease progression, linked with expansion of T-cell precursors, high functional avidity and anti-viral effector function. Broad immune memory pools were narrowed post-infection but NP 105-113 -B*07:02 specific T-cells were maintained 6 months after infection with preserved anti-viral efficacy to the SARS-CoV-2 Victoria strain, as well as new Alpha, Beta and Gamma variants. Our data shows that NP 105-113 -B*07:02 specific T-cell responses associate with mild disease and high anti-viral efficacy, pointing to inclusion for future vaccine design.