Figure 1.

Dynamic Recruitment of FBP17, F-BAR, and N-BAR to Traveling Waves

(A) Diagram of FBP17 versus membrane height change in traveling waves. The lifetime of FBP17 per cycle is about 8 s.

(B) Diagram of constructs used: FBP17S, F-BAR^FBP17(FBP17 aa 1–329), F-BAR^FCHo1(FCHo1 aa 1–275), and N-BAR^Endo (Endophilin-1 aa 1–247).

(C) TIRF micrographs and kymographs of traveling waves of a representative RBL cell co-overexpressing GFP-FBP17S (left) and N-BAR^Endo-mCherry (right). Scale bar, 10 μm.

(D) Left: Intensity profile of GFP-FBP17S (green) and N-BAR^Endo-mCherry (magenta) of the same ~2 × 2-μm² region of interest (ROI) inside the cell in (C). Right: cross-correlation (Xcorr) of the two signals in left shows the phase shift of waves of GFP-FBP17S relative to that of N-BAR^Endo -mCherry. Time resolution: 1.0 s.

(E) Kymographs (left), intensity profile of the ~2 × 2-μm² ROI (middle), and cross-correlation (right) of waves of F-BAR^FCHo1-GFP (green) relative to that of mCherry-FBP17S (magenta). Scale bar, 10 μm. Time resolution: 1.0 s.

(F) Intensity profile (left) and cross-correlation (right) of F-BAR^FBP17_K66E-GFP relative to mCherry-FBP17S. Time resolution: 0.21 s.

(G) Same as (F), but with F-BAR^FBP17_K166A-GFP.

(H) Diagram showing phase shift in waves of F-BAR^FCHo1 and N-BAR^Endo compared with F-BAR^FBP17. Colors denote different protein domains.

(I) Top: Summary diagram of continuous curvature development in traveling waves shown at four exemplary stages. From a flat membrane (t₀ when minimal protein-membrane binding), waves are initiated (t_1, first time point) with shallow curvature, which F-BAR^FCHo1 prefers and binds to, and continuously develop to higher and higher curvature, which attracts F-BAR^FBP17 (t₂) and later N-BAR^Endo (t₃) the most. Plasma membrane is shown as blue mesh. Bottom: 2D cross section of the top. Dashed lines denote the membrane at other time points. Colors denote different protein domains.