Fig. 1.

Loss of Flower does not reduce TfR1 endocytosis but affects LAMP1 endocytosis. A Representative flow cytometric histogram showing TfR1 staining of CTLs activated for five days. TfR1 staining was measured at 0 min, 15 min and 30 min in WT treated with pitstop2, WT and Fwe KO CTLs. The isotype control (upper row) was used for baseline gating. B Quantification of the pooled mean fluorescence intensity of TfR1 internalization in pitstop2-treated, WT and Fwe KO cells. Error bars represent mean ± SEM (N = 3); One-way Analysis of Variance followed by multiple comparison (Dunnett’s), *p < 0.05, ns p > 0.05. C Representative SIM images of LAMP1 internalized in WT and Fwe KO CTLs that were in contact with P815 target cells. CTL and target cells were fixed after 30 min of incubation with anti-LAMP1-647 (magenta) antibody. Scale bar, 2 µm. D Quantification of the fluorescence signal of endocytosed LAMP1 at the immunological synapse (one-third of the cell volume) versus the whole cell in Fwe KO cells in comparison to WT cells. Data given as mean ± SEM (N = 2, n = 34 for both conditions). Data significance was analyzed by Student's t-test, two-tailed ***p < 0.001