Extended Data Fig. 9: Discovery of an N-terminal epitope from SBP.
From: Mapping the T cell repertoire to a complex gut bacterial community
a, Identification of the N-terminal epitope SBP76-84. N-terminal SBP peptides from strains from hCom1d and hCom2d were synthesized and co-cultured with the T cell hybridomas. Truncated peptides were also tested to identify the minimal epitope. 7 of the 13 TCRs were responsive to the synthetic peptides. The remaining 6 were stimulated by C-terminal peptide SBP405-413 as shown in Fig. 4f. There is a strong correlation between the reactivity of TCRs and the sequences of TCR CD3 regions. b, The T cell response to SBP is conserved in different murine settings. We colonized: (1) germ-free C57BL/6 mice with hCom2d by co-housing (horizontal transfer recipient), (2) germ-free C57BL/6 mice with a human fecal community (humanized), (3) germ-free C57BL/6 mice with oral gavage of hCom2d under a condition of a high-fat diet (HFD), and (4) germ-free Swiss Webster mice (SW) with oral gavage of hCom2d. After two weeks, intestinal immune cells were isolated and cocultured with a mix of SBP76-84 and SBP405-413, or PBS as a negative control, using dendritic cells for antigen presentation. Nur77 expression in pTreg or Th17 cells was analyzed by FACS to monitor TCR stimulation (see Supplementary Fig. 1a, c for the gating strategy). SBP-specific T cells were detected in pTreg and Th17 from HT recipient, humanized, and HFD groups, while T cells from SW mice failed to respond to SBP76-84 and SBP405-413. c, We hypothesized that T cells from SW mice recognize a distinct epitope in SBP because of the difference in MHC haplotype between C57BL/6 and SW mice. To test this hypothesis, we co-cultured immune cells from hCom2d-colonized SW mice with a mixture of peptides that tile the whole SBP. We detected SBP-specific pTreg and Th17 cells in hCom2-colonized SW mice. This suggests that SBP-specific T cell induction is preserved across different genetic backgrounds of mice, but—as expected—the epitope recognized is distinct because of the difference in MHC haplotype. p-values were calculated using a two-sided t-test by comparison to PBS treatment as a negative control. *p = 0.05. **p = 0.01. ***p = 0.005. NS > 0.05. Data shown are mean ± standard deviations. n = 8, 8, 14, 8 mice per group from 2 independent experiments (b). n = 15 mice per group from one experiment (c).
