Fig. 8: Perturbation of microvilli shedding affects TCR recovery and T cell proliferation.

a OTI CD8⁺ T blasts were transfected with scrambled or siRNA targeting Vps4a/b. At 24 h post transfection, knock-down efficiency was confirmed by western blot and cells were stimulated on a lipid bilayer presenting pOVA_257-265/H-2K^b/ICAM-1 for 3 h. TISs generation was observed using SEM. Results are representative of three independent experiments. b–d Naïve OTI CD8⁺ T cells transfected with siRNA were stained with CTV and stimulated as in (a). Surface TCRβ recovery was determined by anti-TCRβ-FITC at the indicated time points (b), proliferating populations by Ki-67/NucSpot staining (at 48 h), cell division (c), and the expression of CD25/CD69 (at 24 h) were measured (d). Results are representative of three independent experiments ± SEM. p value were indicated in the figure vs. Scrambled. e–g OTI naive CD8⁺ T cells were pre-stained with TCRβ-Alexa 488 or CTV and stimulated on a lipid bilayer presenting pOVA_257–265/H-2K^b/ICAM-1 in the presence of CK636 (100 μM), JPK (100 nM), or Lat A (237 nM) for 1 h. The number of TCRβ⁺ particles was quantified using CytoFLEX (e). Results are representative of three independent experiments ± SEM. p value was represented vs. DMSO-treated OTI cells stimulated with OVA-H-2K^b + IC1. Cell division and the expression of CD25/CD69 were measured at day 4 (f) and at 24 h (g) post stimulation, respectively. Results are representative of three independent experiments ± SEM. p value was represented in the figure vs. DMSO-treated OTI cells stimulated with OVA-H-2K^b + IC1. Statistics was performed using unpaired two-tailed t test (a, c), one-way ANOVA with post hoc Tukey’s multiple comparisons test (b, e, g). Source data are provided as a Source Data file.