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. 2015 Jan 30;34(6):798–810. doi: 10.15252/embj.201489056

Figure 2.

Figure 2

IP3 receptors are required for dendritic cell fast migration
  1. Relative gene expression of IP3R1 (blue), IP3R2 (red), and IP3R3 (green) in IP3R-silenced DCs. DCs were infected with two lentiviruses encoding for different shRNAs for IP3R isoforms. ShScramble-infected DCs were used as a control. Gene expression was determined by a quantitative PCR. It was calculated with respect to GAPDH expression and normalized to the levels observed in the shScramble. The median plus standard deviation of three independent experiments are shown.
  2. Immunoblot for IP3R type 1 and IP3R type 3 in IP3R-silenced DCs. Actin was used as a control. IP3R3 appears as two bands as previously described (22).
  3. Immunoblot for IP3R type 1 (top) and IP3R type 3 (bottom) in DCs transduced with lentivirus encoding for different IP3R shRNAs. Actin was used as a control.
  4. Relative IP3R2 expression in ShScramble-infected DCs (gray), shIP3R1 (blue), and IP3R3 (green)-silenced DCs. The experiment was performed as described in (A).
  5. Table showing the nomenclature for shRNAs related to their isoform-specific silencing. The shRNAs chosen to pursuit this study are highlighted in red.
  6. Transmigration assay of shScramble-, IP3R(1,3)A (blue)-, IP3R(2,3)B (red)-, and IP3R(1,3)C (green)-expressing DCs. Cells were loaded in the upper chamber of a 5-μm pore collagen-coated transwell assay, recovered from the upper and lower chambers after 16 h, and counted. The median from three independent experiments is shown.
  7. Quantification of the mean cell velocity of shIP3R(1,3)A (blue)-, shIP3R(2,3)B (red)-, and shIP3R(1,3)C (green)-silenced immature DCs migrating in micro-channels. shScramble-infected DCs were used as a control (gray) (n > 100 cells from three independent experiments for shIP3R(1,3)A and shIP3R(1,3)C and two independent experiments for shIP3R(2,3)B). Boxes illustrate 10–90 percentiles of values, and whiskers represent the range of values. P-values were calculated using a Kruskal–Wallis test.
  8. Quantification of the mean cell velocity of shIP3R(1,3)C (green)-silenced immature DCs migrating in micro-channels in the presence of 2 mM BAPTA. shScramble-infected DCs were used as a control (gray) (n > 70 cells from two independent experiments). P-values were calculated using a Kruskal–Wallis test.