Fig. 3: DRAM1 enhances the formation of STX17-SNAP29-VAMP8 complex. | Nature Communications

Fig. 3: DRAM1 enhances the formation of STX17-SNAP29-VAMP8 complex.

From: DRAM1 promotes the stability of lysosomal VAMP8 to enhance autolysosome formation and facilitates the extravasation

Fig. 3

A Levels of GFP-SNAP29 and GFP-VAMP8 co-immunoprecipitated by Flag-STX17 in Control and DRAM1 OE cells. 293 T cells transfected with the indicated constructs were immunoprecipitated using anti-Flag antibodies and subjected to WB analysis. B Quantification of relative levels of GFP-SNAP29 and GFP-VAMP8 that are co-immunoprecipitated by Flag-STX17 in Control and DRAM1 OE cells. The level of co-precipitated GFP-SNAP29 and GFP-VAMP8 were quantified by Image J and normalized to the corresponding Flag-STX17 (set to 1 in control cells). C, D Colocalization of Flag-STX17 with VAMP8 in Control or DRAM1 OE HeLa cells. Representative confocal images are shown, and the percentage intensity of STX17 colocalized with VAMP8 was quantified by Image J (n = 9 cells/group). Data are shown as means ± SD representative of two independent experiments. Scale bars: 10 μm. E Levels of endogenous SNAP29 and VAMP8 co-immunoprecipitated by STX17 in Control and DRAM1 OE 293 T cells. Cells transfected with the indicated constructs were immunoprecipitated using anti-STX17 antibodies and subjected to WB analysis. F Quantification of relative levels of SNAP29 and VAMP8 that are co-immunoprecipitated by STX17 in Control and DRAM1 OE cells. The level of co-precipitated SNAP29 and VAMP8 were quantified by Image J and normalized to the corresponding STX17 (set to 1 in control cells). G Levels of endogenous STX17 and VAMP8 immunoprecipitated in Control and DRAM1 OE cells. H Quantification of relative levels of STX17 and VAMP8 that are co-immunoprecipitated by SNAP29 in Control and DRAM1 OE 293 T cells. I Levels of endogenous STX17 and SNAP29 immunoprecipitated in Control and DRAM1 OE cells. 293 T cell extracts were immunoprecipitated with anti-VAMP8 and the resulting precipitants were analyzed by WB using the indicated antibodies. J Quantification of relative levels of STX17 and SNAP29 that are co-immunoprecipitated by VAMP8. Data in (B, F, H, J) are presented as mean ± SD, n = 3 independent experiments. Data in (A, E, G, I, C) are representative of three and two independent experiments, respectively. P values were calculated by two-tailed Student’s t tests (B, D, F, H, J). P values are indicated in the figure.

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