Fig. 3: Kinesin-1 regulates Ag degradation and acidification in the endocytic pathway. | Nature Communications

Fig. 3: Kinesin-1 regulates Ag degradation and acidification in the endocytic pathway.

From: Kinesin-1 regulates antigen cross-presentation through the scission of tubulations from early endosomes in dendritic cells

Fig. 3

a CD8α+, CD11b+ DCs and BMDCs from WT (blue histogram) or cKOKif5b (red histogram) mice were incubated for 10, 30 or 60 min at 37 °C or for 10 min at 4 °C with Alexa-Flour-647-OVA prior to flow cytometry analysis. The figure shows a histogram that was representative of three independent experiments. b CD8α+, CD11b+ DCs and BMDCs from WT (blue line) or cKOKif5b (red line) mice were incubated for 30 min at 37 °C with DQ-OVA. Next, the cells were washed. DQ-OVA degradation was analysed by flow cytometry after different chase periods. Graphs are representative of four independent experiments. Statistical significance was determined by the two-way ANOVA and Sidak test’s correction for multiple comparison. ***P < 0.0001. c Degradation of DQ-OVA in BMDCs from WT (DMSO blue line, Conc-A or Cath I purple line) or cKOKif5b (DMSO orange line, Conc-A or Cath I yellow line) mice pretreated or not with concanamycin A (Conc-A) or a cathepsin inhibitor-I (Cath I). Graphs are representative of four independent experiments. Statistical significance was determined by the two-way ANOVA and Sidak test’s correction for multiple comparison. **P < 0.005; ***P < 0.0001. d The pH values in CD8α+, CD11b+ DCs and BMDCs from WT (blue line) or cKOKif5b (red line) mice were determined by FACS after a pulse and different chase periods. Graphs are representative of four independent experiments. Statistical significance was determined by the two-way ANOVA and Sidak test’s correction for multiple comparison. *P < 0.05; **P < 0.005; ***P < 0.0001. e Protease activity of CatB/L in early (20 min) or late (120 min) using total cell lysate from BMDCs from WT (blue histogram) or cKOKif5b (red histogram) mice was measured with a specific fluorescent substrate. Graphs are representative of three independent experiments. Statistical significance was determined by the two-way ANOVA and Sidak test’s correction for multiple comparison. ***P < 0.0001. a–e Graphs show mean ± S.E.M.

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