Fig. 3: SEMA3A negatively regulates CD8⁺ T cell adhesion, motility and migration through NRP1.

a Representative brightfield and IRM images of stimulated OT-I T cells adhering to ICAM-1/CXCL12 coated plates with either SEMA3A_S-P or IgG. Scale bar = 20 μm. b Contact area per single OT-I T cell. Representative of three independent experiments. c Frequency of cell polarity from brightfield images. A polarity of 1 indicates a shape of a perfect circle, 0 a rectangular shape. Representative images of OT-I T cells illustrated above graph. d Spider plots showing the migration paths of T cells pre-treated with either NRP1 blocking antibody or isotype control antibody on similar plates as in (a). Scale bar = 100 μm. e Single cell distance (left) and single cell velocity (right) from same experiment as (d) (n = 380 cells for IgG and isotype control, 314 cells for IgG and anti-NRP1, 744 cells for Sema3A and isotype control, anf 403 for Sema3A and anti-NRP1). f Spider plots showing the migration path of OT-I T cells on ICAM-1/CXCL12 coated flow cells as in (a), with flow rates at 0 or 80 μm/s. Arrows indicate flow direction. Scale bar = 50 μm. g Percent of OT-I cells that detach in same experiment as (f) (n = 19 cells with control IgG and 73 cells with Sema3A). Representative of two independent experiments. h Number of stimulated OT-I T cells from individual mice (n = 3) able to transmigrate through 3 μm Boyden chamber in indicated conditions. OT-I T cells were pre-treated with either a blocking NRP1 antibody or isotype control antibody. Data representative of two independent experiments. Abbreviations: ns, not significant. Error bars are means ± SEM (b, g) or SD (e, h) from representative experiments (b, h) or combined from 2 (g) and 5 (e) independent experiments. *P = 0.04 (g), **P = 0.003 (g), ****P < 0.0001 (b, e, g, h) by two-tailed Student’s t-test (b), two-way ANOVA (e, g) or one-way ANOVA (h). Source data are provided as a Source Data file.