TCR clonotype analysis and single-cell transcriptomics distinguish antigen-specific human memory CD4+ T cells which recognize macrophages infected with Mycobacterium tuberculosis 2481

Abstract Description CD4+ T cell responses to Mycobacterium tuberculosis (Mtb) antigens processed and presented by infected macrophages leads to control of infection. However, not all T cells specific for Mtb antigens recognize infected macrophages. Using ex vivo co-culture of primary human T cells obtained from healthy individuals with latent Mtb infection (LTBI) together with autologous macrophages infected with Mtb, we estimate that >80% of memory CD4+ T cells recognize Mtb-infected macrophages. Using TCR sequencing and GLIPH2 grouping of activated CD4+ T cells, we identified “likely” and “known” Mtb antigen-specific TCR clonotypes based on previous TCR annotation. While 70-80% of unique and >90% of total TCR clonotypes found in circulation were linked to recognition of infected macrophages, a subpopulation 10-20% were not. Using single-cell transcriptomics, TCRs were mapped to function. IFNg, TNF, and lymphotoxin-alpha (LTa) were preferentially expressed only among clusters containing clonally-expanded TCRs. While top GLIPH2 groups mapped to several clusters, those co-expressing IFNg, TNF, LTa, IL-2, and GM-CSF were preferentially enriched for Mtb-specific TCRs. GM-CSF and “polyfunctional” T cells have been shown to play protective roles against Mtb infection. These results link the effector functions of antigen-specific memory CD4+ T cells to recognition of the infected macrophage and stable LTBI. T cell recognition assays would enhance the evaluation and development of new TB vaccines. Funding Sources NIH K08 AI163407 (Carpenter) NIH R21 AI167675 (Carpenter, Freeman) American Lung Association Catalyst Award 924107 (Carpenter) University Hospitals Research Funds (Carpenter) NIH 75N93019C00071 / HI-IMPAcTB (Fortune / Flynn / Boom) Topic Categories Microbial, Parasitic, and Fungal Immunology (MPF)