Fig. 8: SKW-3-CD3+ and SKW-CD3+CD8+ cell lines expressing age-specific TCRs. | Nature Immunology

Fig. 8: SKW-3-CD3+ and SKW-CD3+CD8+ cell lines expressing age-specific TCRs.

From: Newborn and child-like molecular signatures in older adults stem from TCR shifts across human lifespan

Fig. 8

a, Selection of A2/M158+CD8+ TCR specifically identified in certain age groups. p, proliferation; aa, amino acid. b,c, Representative A2/M158 tetramer-PE staining of SKW-3-CD3+ (b) and SKW-3-CD3+CD8+ (c) TCR-expressing cell lines. d, Median MFI A2/M158 tetramer-PE of SKW-3-CD3+ (open bars) and SKW-3-CD3+CD8+ TCR-expressing cell lines (closed bars) (n = 5 independent experiments, median and interquartile range (IQR)); dotted line indicates MFI threshold set by the parental cell line expressing no TCR. MFI, median fluorescence intensity. e,f, Representative staining (e) and median MFI (f) of A2/M158 tetramer-PE staining with a normal CD8-binding site (A2/M158-WT, lightly shaded), knockout CD8-binding site (A2/M158-KO open) and enhanced CD8-binding site (A2/M158-Enh, closed) tetramer of SKW-3-CD3+CD8+ TCR-expressing cell lines (n = 2 independent experiments); dotted line indicates MFI threshold set by the parental cell line expressing no TCR. g, Representative CD69-PECy7 expression following peptide titration for public and age group specific TCRs expressed in SKW-3-CD3+ cell lines. h, Percentage of maximum CD69-PECy7 MFI for age-specific TCRs expressed on SKW-3-CD3+ (open circles) and SKW-3-CD3+CD8+ (closed circles) cell lines following peptide titration; EC50 indicated by the dotted line (n = 3 independent experiments, median and range). i, Representative CD69-PECy7 expression profiles of SKW-3-CD3+ TCR-expressing cell lines following stimulation with the M158–66 peptide with single alanine substitutions, except for binding site (p2). j, Percentage of maximum CD69-PECy7 MFI following stimulation with M158–66 alanine scan peptides for age-specific TCRs expressed on SKW-3-CD3+ cells (n = 3 independent experiments, median and range). Statistical analysis was performed using a two-sided Kruskal–Wallis with Dunn’s correction for multiple tests between the public TCR and the age-specific TCRs within the same cell line (d,j, black) or between the same TCRs across two cell lines (d, blue). P values are indicated above the graphs.

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