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

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.