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. 2020 Dec 7;24(1):101900. doi: 10.1016/j.isci.2020.101900

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© 2020 The Author(s)

This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

PMC Copyright notice

NLRC5 favors the activation of Vγ9Vδ2 T cells

(A) Polyclonal Raji cells were transduced with NLRC5-coding or mock vector and treated with zoledronate (+ZOL) for 24 h (bottom panel) or left untreated (top panel). BTN3A1-3 surface expression was analyzed by flow cytometry. Graphs depict a quantification of BTN3A1-3 geometric MFI (gMFI; top and bottom left), and the histogram overlays (bottom right) show BTN3A1-3 expression for unstained (US; black, gray), mock- (blue) and NLRC5-transfected polyclonal Raji cells (pink). Results represent six independent measurements.

(B) Specific killing of NLRC5- (pink) or mock-transduced polyclonal Raji cells (blue) was measured after 24 h of co-culture with V_γ9V_δ2 T cells at 3 effector-to-target ratios (1:1, 5:1, and 10:1) in the presence of ZOL. Depicted are the results for three independent healthy donors (HD) as mean ± SD (n = 3 technical replicates). The results are representative of four independent healthy donors and three independent experiments.

(C) BTN2A1 and BTN3A1, A2, and A3 mRNA levels (relative to POLR2A mRNA) were assessed by qRT-PCR in the NLRC5- (red and violet) or mock-transduced Raji subclones (blue). The results depict mean ± SD (n = 3 technical replicates) and are representative of 2 independent experiments.

(D) Specific killing of the NLRC5- or mock-transduced Raji subclones was measured after 24 h of co-culture with V_γ9V_δ2 T cells at 3 effector-to-target ratios in the presence of ZOL. n.d. correspond to conditions where no specific lysis of the target cells by the V_γ9V_δ2 T cells was detected. Depicted are the results for three independent healthy donors (HD) as mean ± SD (n = 3 technical replicates). The results are representative of six independent healthy donors and three independent experiments. (A–D) Statistical differences between the condition with and without NLRC5 overexpression were calculated using paired Student's t test (A) or post hoc Student's t tests adjusted for multiple comparisons using the Holm-Sidak method (B–D). ∗p ≤ 0.05, ∗∗p ≤ 0.01, ∗∗∗p ≤ 0.001. Only statistically significant differences are illustrated.