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. 2020 Oct 20;23(11):101709. doi: 10.1016/j.isci.2020.101709

Figure 1.

Figure 1

Deletion of FcRγ in NK Cells Leads to Enhanced Cytokine Production in Response to CD16 Cross-linking

(A) Contour plots show a mixture of NK (CD3−CD56+), T (CD3+CD56- and CD3+CD56+), and non-NK/non-T (CD3−CD56-) cells after expansion of PBMC samples enriched for NK cells. NK cells (gated, red) were further analyzed for FcRγ expression following nucleofection with FCER1G-targeting Cas9/gRNA ribonucleoprotein (FCER1G-RNP) or non-targeting Cas9/gRNA ribonucleoprotein (ctrl RNP) after 10 days of in vitro culture. Data are representative of 5 independent experiments performed on NK cells from 10 donors.

(B) Time course analysis of FcRγ depletion in FCERIG-RNP-treated NK cells (red), with CD3+CD56- T cells (blue) overlaid for comparison. The staining pattern obtained using a matched isotype control antibody (iso ctrl) is shown.

(C–E) Contour plots show flow cytometric analysis of (C) TNF-α, (D) IFN-γ production, and (E) cell surface CD107a expression by FcRγ+ and FcRγ− NK subsets from a representative donor without (−) or with (+) CD16 cross-linking (CXL). Numbers represent the relative percentages of FcRγ+ or FcRγ− subset that produced indicated cytokine or displayed CD107a. Line graphs show the relative percentages of FcRγ+ or FcRγ− subset that produced indicated cytokine or displayed CD107a from several donors in response to CD16 CXL (n = 10). Circles connected by a line designate data obtained from the same donor sample.

(F–H) Line graphs show the relative percentages of conventional NK (cNK) cells and g-NK cells that produced (F) TNF-α, (G) IFN-γ or (H) displayed CD107a following CD16 CXL (n = 7). Wilcoxon signed-rank test; ∗p < 0.05, ∗∗p < 0.01, n.s., not significant.