Fig. 6: MdSYP121 interacts with MdSNAP33 and forms a SNARE complex.
a Transient expression of a 35S::MdSNAP33-GFP fusion construct in N. benthamiana. Green fluorescence was observed after transient infection for 4 days using a confocal microscope. Bars = 10 μm. In vivo interactions between MdSYP121 and MdSNAP33 were determined using BiFC. C-terminal and N-terminal fragments of YFP (indicated in the figure as YFPC and YFPN, respectively) were fused to the C-terminus of MdSYP121 and MdSNAP33, respectively. Combinations of YFPN and YFPC with the corresponding MdSYP121 and MdSNAP33 constructs were used as negative controls. Fluorescence of YFP represents protein-protein interactions. Yellow fluorescence was observed using a confocal microscope after transient infection for 4 days. Bars = 10 μm. b GST pull-down assays were performed using HA antibody. Western blot analysis results showing protein expression levels. c MdSYP121 together with MdSNAP33 forms SDS-resistant SNARE complexes. SDS-resistant MdSYP121-containing complexes are detectable in Vec and MdSYP121-OE calli lines. Proteins were extracted from Vec and MdSYP121-OE lines and incubated together with GST-MdSNAP33 protein. The proteins were loaded on a 10% polyacrylamide gel and subjected to immunoblot analysis in conjunction with anti-GST. The lower panel shows the same immunoblot protein samples that were detected with anti-HA
