Mtb ligands are recognized by human Mtb-reactive CD8⁺ T cells.

<p>CD8⁺ T cells from four healthy, eight LTBI, and four Mtb-infected donors were incubated in an IFN-γ ELISPOT plate with autologous DC pulsed with each of the 28 validated Mtb peptides. Shown are the IFN-γ spot forming units for the 12 peptides for which at least one donor responded.</p

Mtb infection alters the HLA-E host ligand repertoire and results in a minor Mtb-specific ligand pool.

<p>Percentage of total ion intensity as determined in the materials and methods section of indicated HLA-E ligand groups from uninfected (A) or Mtb infected cells (B).</p

Production of soluble HLA-E from Mtb-infected cells.

<p>A) U373 or DC were infected with the H37Rv strain of Mtb (multiplicity of infection (MOI):30, 10, or 3) for 18 hours, then used as antigen presenting cells in an ELISPOT assay with IFN-γ production by D160 1–23, an HLA-E restricted CD8⁺ T cell clone, as the readout. B) U373 cells expressing sHLA-E, and grown to high density in a hollow-fiber bioreactor, were infected with Mtb. sHLA-E was measured in the cell supernatant using ELISA.</p

Mtb peptides associated with sHLA-E.

<p>Mtb peptides associated with sHLA-E.</p

An Mtb ligand derived from Rv0634A is eluted from sHLA-E molecules.

<p>A) Extracted ion chromatogram of ion 658.8210 (EIEVDDDLIQK) in Mtb infected ligand pool (blue trace) and synthetic peptide (black trace). B) Annotated MS2 fragment ion spectra for the identified Mtb peptide EIEVDDDLIQK in Mtb infected ligand pool (blue trace) and synthetic peptide (black trace).</p

The CD8⁺ T cell response to Rv0634A_19-29 is restricted by HLA-E.

<p>A) A549 cells were pulsed with the Rv0634_19-29 peptide and used as antigen presenting cells in an ELISPOT assay with IFN-γ production by CD8⁺ T cells from mismatched donors as a readout. B-D) D481 CFSE-labeled CD8+ T cells were isolated and cultured with Rv0634A_19-29 peptide-pulsed autologous macrophages and DC. CFSE-dim cells were sorted and stained with the Rv0634A_19-29 HLA-E tetramer and antibodies against CD3 and CD8. As a control, the D481 line was also stained with a non-HLA-E tetramer (C), or with anti-NKG2A (D) do demonstrate that the tetramer staining is specific. Whole PBMC were stained with the anti-NKG2A antibody as a positive control. E) The mismatched A549 cell line was pulsed with the Rv0634A_19-29 peptide and used as antigen presenting cells with D481 CD8⁺ T cell line.</p

Class I leader peptides eluted from sHLA-E molecules.

<p>A) Representative annotated MS2 fragment ion spectra for the identified leader peptides from HLA-A*02:01, HLA-B*18:01, and HLA-C*05:01 that were eluted from purified sHLA-E. B) Length distribution of all non-redundant HLA-E ligands identified combined from Mtb infected and uninfected cells. C) Peptide binding motif of nonamer peptide sequences eluted from HLA-E. Motif was generated using Seq2Logo.</p

Rhesus Macaque TRAJ genes

TRAJ genes extracted from the rhesus macaque genome. The gene features reported include J-NONAMER, J-SPACER, and J-HEPTAMER. The FASTA header fields include the species name, rhesus macaque chromosome 7 accession number (NC_007864.1), start and end positions of the gene feature in the rhesus macaque chromosome 7, strand (-1 indicates that the reported sequences are the reverse complement), gene name, and gene feature

Rhesus Macaque TRAV and TRDV genes

TRAV and TRDV genes extracted from the rhesus macaque genome. The gene features reported include L-PART1, V-INTRON, V-EXON, V-HEPTAMER, V-SPACER, and V-NONAMER. The FASTA header fields include the species name, rhesus macaque chromosome 7 accession number (NC_007864.1), start and end positions of the gene feature in the rhesus macaque chromosome 7, strand (-1 indicates that the reported sequences are the reverse complement), gene name, and gene feature