Figure 5.
FCRL3 expression is causally associated with reduced T cell activation. (A) Expression of FCRL3, TOX, and EOMES transcripts was measured in FCRL3-KO and control cells by RT-qPCR. Transcript expression was normalized to the housekeeping gene UBE2D2 and is shown relative to control samples. N = 5 independent experiments; paired t test, two-tailed. From left to right: ****P < 0.0001, *P = 0.0475, **P = 0.0040. (B) Surface expression of FCRL3 in Jurkat T cells transduced with FCRL3-lentivirus or empty lentivirus as a control. One representative experiment of N = 4 is shown. (C and D) Surface CD25 expression in FCRL3-transduced Jurkat cells. Cells were stimulated for 24 h with anti-CD3/CD28-coated beads. One representative experiment is shown in C, while the bar graphs in D show the compiled results of N = 4 independent experiments. The gMFI and percentage of CD25+ cells are both shown. Mean ± SEM; paired t test, two-tailed. From left to right: **P = 0.0052, **P = 0.0033. (E) Schematic representation of the EGFR-FCRL3 chimeric proteins. (F) Western blot showing the expression of the different truncated chimeric proteins in Jurkat T cells, using an anti-EGFR antibody. Data are representative of N = 2 experiments. (G) Surface expression of the EGFR in Jurkat T cells transduced to ectopically express the EGFR-FCRL3 chimeras. Data are representative of N = 4 independent experiments. (H) Surface expression of CD25 in Jurkat T cells expressing the EGFR-FCRL3 chimeras. Cells were stimulated for 24 h with anti-CD3/CD28-coated beads. One representative experiment is shown. (I) Surface expression of CD25 in stimulated (24 h) Jurkat T cells expressing the EGFR-FCRL3 chimeras. The compiled results of N = 4 independent experiments as in E are shown. Mean ± SEM; paired t test, two-tailed. Left panel, from top to bottom: ***P = 0.0007, *P = 0.0151, **P = 0.0046, ***P = 0.0002, **P = 0.0030, **P = 0.0013. Right panel, from top to bottom: ***P = 0.0001, n.s. (not significant), P = 0.1005, *P = 0.0326, **P = 0.0014, **P = 0.0050, *P = 0.0157. Data underlying this figure can be found in Data S4. Source data are available for this figure: SourceData F5.

FCRL3 expression is causally associated with reduced T cell activation. (A) Expression of FCRL3, TOX, and EOMES transcripts was measured in FCRL3-KO and control cells by RT-qPCR. Transcript expression was normalized to the housekeeping gene UBE2D2 and is shown relative to control samples. N = 5 independent experiments; paired t test, two-tailed. From left to right: ****P < 0.0001, *P = 0.0475, **P = 0.0040. (B) Surface expression of FCRL3 in Jurkat T cells transduced with FCRL3-lentivirus or empty lentivirus as a control. One representative experiment of N = 4 is shown. (C and D) Surface CD25 expression in FCRL3-transduced Jurkat cells. Cells were stimulated for 24 h with anti-CD3/CD28-coated beads. One representative experiment is shown in C, while the bar graphs in D show the compiled results of N = 4 independent experiments. The gMFI and percentage of CD25+ cells are both shown. Mean ± SEM; paired t test, two-tailed. From left to right: **P = 0.0052, **P = 0.0033. (E) Schematic representation of the EGFR-FCRL3 chimeric proteins. (F) Western blot showing the expression of the different truncated chimeric proteins in Jurkat T cells, using an anti-EGFR antibody. Data are representative of N = 2 experiments. (G) Surface expression of the EGFR in Jurkat T cells transduced to ectopically express the EGFR-FCRL3 chimeras. Data are representative of N = 4 independent experiments. (H) Surface expression of CD25 in Jurkat T cells expressing the EGFR-FCRL3 chimeras. Cells were stimulated for 24 h with anti-CD3/CD28-coated beads. One representative experiment is shown. (I) Surface expression of CD25 in stimulated (24 h) Jurkat T cells expressing the EGFR-FCRL3 chimeras. The compiled results of N = 4 independent experiments as in E are shown. Mean ± SEM; paired t test, two-tailed. Left panel, from top to bottom: ***P = 0.0007, *P = 0.0151, **P = 0.0046, ***P = 0.0002, **P = 0.0030, **P = 0.0013. Right panel, from top to bottom: ***P = 0.0001, n.s. (not significant), P = 0.1005, *P = 0.0326, **P = 0.0014, **P = 0.0050, *P = 0.0157. Data underlying this figure can be found in Data S4. Source data are available for this figure: SourceData F5.

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