Fig 2: Identification of adaptive NK cells from a 24-year-old female donor. a The expression of KLRC2 in each donor cells within the “Adaptive NK” cluster were shown in violin plots. The y-axis represents log-normalized expression value. b The t-SNE plot demonstrates four distinct NK clusters from BM of the 24-year-old female donor. c The expression of KLRC2 of the 24-year-old female sample was overlaid on the t-SNE plot. d Violin plots demonstrate the expression of FCER1G, ZBTB16, SH2D1B, and SYK in four clusters from the 24-year-old female donor sample. The y-axis represents log-normalized expression value. e Violin plots demonstrate the expression of CD3E, CD3D, and CD3G in four clusters from the 24-year-old female donor sample. The y-axis represents log-normalized expression value. f Expression of NKG2C on NK cells (gated on PI-Lin-TCRa/ß-TCR?/d-CD56+ cells) was assessed by flow cytometry using cryopreserved samples from four donors. g Selected gene sets enriched in the “Adaptive NK” cluster compared to the rest of the cells were plotted. See also Supplementary Fig. 6

Fig 3: FceRI?-NKG2C+CD57+ cells markedly expand in HCMV-seropositive individuals and tightly correlate with individual FceRI?- memory pool size. (A) Gating strategy for the identification of peripheral blood FceRI?- and FceRI?+ CD16+CD56dim NK cell subsets. Numbers represent percentage of gated cells. (B,C) Stratification of CD16+CD56dim NK cells according to NKG2C and CD57 expression, within FceRI?- (?-, grey) and FceRI?+ (?+, empty) cell pools, of 154 HCMV-seropositive (HCMV+) (B) and 75 HCMV-seronegative (HCMV-) (C) healthy individuals. Bars represent median with interquartile range. p values of pairwise comparisons were calculated with Wilcoxon (FceRI?- vs. FceRI?+) and Mann–Whitney (HCMV+ vs. HCMV-) non-parametric tests. (*) symbol indicates statistical comparison with corresponding FceRI?+ subset (FceRI?- vs. FceRI?+): (*) <0.05, (**) <0.005, (***) < 0.0001, (****) < 0.00001; (#) symbol indicates statistical comparison with corresponding subset of HCMV- donors (HCMV+ vs. HCMV-): (#) < 0.05, (##) < 0.005, (####) < 1 × 10-7. (D) Correlation between the percentage of FceRI?-NKG2C+CD57+ (grey, upper panels) or FceRI?+NKG2C+CD57+ (white, lower panels) subsets with the size of the FceRI?- memory NK cell pool in HCMV+ (left panels) and HCMV- (right panels) individuals. Spearman’s correlation coefficients (r) and curve slopes are reported when 2-tailed p values were significant (<0.05): (****) < 1 × 10-7.

Fig 4: Anti-HCMV IgG levels impact FceRI?- memory NK cell pool abundance and correlate with the expansion of FceRI?-NKG2C+CD57+ subset in seropositive individuals. (A) Percentage of FceRI?- memory (within CD16+CD56dim) NK cells in HCMV+ healthy donors stratified for higher (>10 arbitrary units, au, grey triangles) or lower (=10 au, empty triangles) levels of anti-HCMV antibodies. Relative percentage of FceRI?- (?-, grey circles) and FceRI?+ (?+, empty circles) subsets in individuals with >10 au (B) or =10 au (C) anti-HCMV antibodies. Bars represent median with interquartile range. p values of pairwise comparisons were calculated with Wilcoxon and Mann–Whitney (>10 au vs. =10 au) non-parametric tests, as appropriate. (*) symbol indicates statistical comparison with corresponding FceRI?+ subset (FceRI?- vs. FceRI?+): (*) < 0.05, (**) = 0.01, (***) < 0.005, (****) < 0.001, (*****) < 0.00001; (#) symbol indicates statistical comparison with corresponding subset of individuals producing lower levels of anti-HCMV antibodies (>10 au vs. =10 au): (#) < 0.05, (##) < 0.01, (###) < 0.0005. (D) Correlation between the relative abundance of FceRI?- (left panel) or FceRI?+ (right panel) NKG2C+CD57+ cells and anti-HCMV antibody concentration. Individuals possessing a larger (higher the median value, grey diamonds) or smaller (lower than median value, empty diamonds) given subset were analysed separately. Spearman’s correlation coefficients (r) and curve slopes are reported when 2-tailed p values were significant (<0.05): (*) < 0.05, (***) < 0.005.

Fig 5: FcεRIγ−NKG2C+CD57+ subset enhanced in vitro expansion driven by ITP patient-derived platelets. Expansion index, expressed as the cell number fold increase in the presence of healthy donor’s (CTR) or ITP patient’s (ITP) platelets (plts) over that attained in the presence of Raji cells alone, of FcεRIγ− and FcεRIγ+ NK cells (A), or after stratification according to NKG2C and CD57 expression in FcεRIγ− (B) (upper panel) and FcεRIγ+ (B) (lower panel) NK cell pools. (C) Plots of a representative HCMV+ donor cultured with either CTR or ITP platelets. Numbers represent percentage of gated cells. Data are representative of 12 independent experiments. p values of pairwise comparisons were calculated by Wilcoxon non-parametric test and reported when significant (<0.05): (***) p < 0.005.