Figure 3.

In situ FLIM-FRET identifies VAMP7 as a heterodimer-dependent interaction partner of Stx17. a and b, intensity images and their corresponding fluorescence lifetime maps probing changes in lifetime and proximity between donor-only or donor and acceptor samples in rapamycin-treated cells. Scale bars, 10 μm. Shown is EGFP-VAMP8 (a) and EGFP-VAMP7 (b) expressed alone or co-expressed with either WT mCherry-Stx17 or mCherry-Stx17[Q196G]. c and d, mean single-pixel FRET efficiency histograms, enveloped by S.E., derived from the data sets presented in a and b, respectively. Statistical significance was tested using a Mann–Whitney test to compare integral values > 0.1 (n = 7–10 cells). e, schematic demonstrating the differing R-SNARE compositions, which have an unknown impact on complex structure and fluorophore separation distance. ns, nonsignificant (p ≥ 0.05); **, p = 0.0005–0.005.