Figure 2.

The effects of ER-SNARE overexpression on secretion and retrograde Golgi-ER transport in tobacco protoplasts. Protoplasts were co-electroporated with a constant amount of plasmid DNA encoding for α-amylase and increasing amounts of plasmid DNA encoding for full-length wtUSE1 (A), SYP71-GFP [(B) light gray], or SYP72-GFP [(B) dark gray]. SYP71-GFP [(C) light gray], and SYP72-GFP [(C) dark gray], respectively, were also coexpressed with α-amylase-HDEL which is normally retained in the ER by efficient COPI-mediated retrograde transport from the Golgi. Secreted and protoplast retained levels of α-amylase activity were measured and the secretion index calculated as described in Materials and Methods. The first column shows as a control the expression of the reporter alone, the other columns the coexpression with constructs in different concentrations as indicated below the axis. USE1 inhibits secretion (A) while SYP71, SYP72 do not (B). Expression of either SYP71-GFP or SYP72-GFP lowers the secretions index of α-amylase-HDEL indicating a more efficient retrograde Golgi-ER transport (C). Total activity for SYP71-GFP [(D) light gray] or SYP72-GFP [(D) dark gray] decreased only at very high amounts of electroporated DNA. Standard deviations are given as vertical lines in each column.