DC F-actin depolymerization affects immune synapse structure and T cell priming efficiency . (A) Bright field images of mature DCs treated with 1 µM MycB (right) or DMSO (left). Scale bar: 10 µm. (B) Flow cytometry profiles of DMSO and 1 µM MycB-treated mature DCs. Cd11c/MHC II plots are pregated on FSC-A/SSC-A population defined by black oval on the left, representative example of three biological replicates. (C) Immunofluorescence images of synapses formed between DMSO or 1 µM MycB-treated mature DCs and T cells. Upper panel: overview pictures, yellow dotted line outlines DC. Scale bar: 5 µm. Middle panel: en face view on the synaptic interface. Scale bar: 1 µm. Lower panel: surface reconstruction of the synaptic interface. (D) Percentages of mono- and multifocal synapses formed between T cells and 1 µM MycB-treated mature DCs, n = 20 cell duplets for each condition, three biological replicates, mean ± SD. (E) Percentages of activated T cells assessed by CD62L/CD69 surface expression after 16 h of coculture with DMSO or 1 µM MycB-treated mature DCs at indicated OVA_323-339 peptide concentrations, three biological replicates, mean ± SD. (F) CFSE dilution profile of T cells after 96 h of coculture with DMSO or 1 µM MycB-treated mature DCs at indicated OVA_323-339 peptide concentrations, representative example of three biological replicates. (G) Proliferation indices of CFSE-labeled T cells after 96 h of coculture with DMSO or 1 µM MycB-treated mature DCs at indicated OVA_323-339 peptide concentrations, three biological replicates, mean ± SD. (H) Absolute T cell numbers after 96 h of coincubation with DMSO or 1 µM MycB-treated mature DCs at indicated OVA_323-339 peptide concentrations, three biological replicates, mean ± SD. Data in E, G, and H were tested for normal distribution, transformed if necessary, and tested by using Student’s t test. FSC-A, forward scatter–A; ns, not significant; SSC-A, side scatter–A. *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001.