TCRs from GLIPH2 groups are specific for Mtb antigens. (A–C) (A) Bar graphs of CD69 expression by flow cytometry (gated on CD4⁺ TCRβ⁺ Live-Dead^Lo) for SKW-3 cells transduced with a TCR from indicated GLIPH2 groups 18 h after coculture with autologous B cells loaded with MTB300 megapool, 20-peptide subpools, or (B) with individual peptides, DMSO, or after treatment with αCD3/CD28 mAb-coated beads for R%SGGEAKNI and (C) RKQG%E TCR-transduced SKW-3 cells. (D) Representative flow plots of CD69 and TCRβ expression gated on total CD4⁺ SKW-3 cells after transduction with a TCR expressing the S%SGTKYNE motif in response to cognate peptide (top) or irrelevant peptide (bottom). (E–G) Bar graphs of CD69 expression (of CD4⁺ TCRβ⁺) for (E) the S%SGTKYNE TCR, (F) the SSPGQGG%NYG TCR, and (G) the %MPE TCR in response to individual peptides. (H) Single values for each condition are plotted, representing two to three independent experiments for (H) each cloned TCR and respective GLIPH2 group. Cognate peptides and GLIPH2-predicted HLA restrictions are listed. (I) Sequence logo plots show the probability of each amino acid for CDR3β (top) and CDR3α (bottom) motifs for each GLIPH2 group containing TCRs where Mtb antigen specificity was established. Created using WebLogo3. Numbers of CDR3 sequences used for each plot are indicated (top-right). * indicates two, and ** indicates three different CDR3α lengths and a lack of CDR3α consensus within the GLIPH2 group.