Fig. 5.

Syt VII-YFP translocates from peripheral domains of lysosomal compartments to the plasma membrane during DC maturation. (A) Immature DCs were transduced with Syt VII-YFP (green) and loaded with Texas red (TR) dextran (red) prior to fixation. Images are 0.8 μm optical slices from the middle and top of a confocal Z-stack. Arrows point to peripheral domains of dextran loaded lysosomes containing Syt VII-YFP. (B) Live confocal image of a maturing (1 h LPS) DC loaded with TR-dextran (red) and transduced with MHC-II-GFP (green). (C) Live confocal images of DCs transduced with either MHC-II-GFP (green) [left] or Syt VII-YFP (green) [middle], loaded with TR-dextran (red), and induced to mature for 24 h with LPS/cluster disruption. Right panel: FACS profile comparing surface levels of endogenous Syt VII in immature DCs, or DCs induced to mature for 24 h with LPS/cluster disruption. Anti-Syt VII NH₂-terminal antibodies were added at 4 °C to detect surface exposed Syt VII. The background control (black) represents secondary antibody alone. About 36% of LPS-treated cells had surface levels of Syt VII significantly above that of immature cells. The results shown are representative of several independent assays. Bars = 5 μm.