Figure 6: The N-BAR domain is responsible for PI clustering. | Nature Communications

Figure 6: The N-BAR domain is responsible for PI clustering.

From: BIN1/M-Amphiphysin2 induces clustering of phosphoinositides to recruit its downstream partner dynamin

Figure 6

(a) Binding of the PtdIns(4,5)P2-binding motif PH(PLCδ) (in cyan) on flat membranes containing 5% PtdIns(4,5)P2 (0.1% GloPIP2, in red) previously incubated with 2 μM of the BIN1 N-BAR or 2 μM of the BIN1-mutated N-BAR D151N. Red arrows point to the formation of membrane tubes after N-BAR incubation. (b) Binding of the PtdIns(4,5)P2 motif PH(PLCδ) to flat membranes containing 5% PtdIns(4,5)P2 in control conditions (grey) and in the presence of the N-BAR domain (magenta) or the N-BAR D151N (orange). (c,d) HeLa cells co-expressing GFP-BIN1 SH3-ΔC (c) or GFP-BIN1 D151N (d) mutants along with mCherry-PH(PLCδ) (see Methods). (e,f) Live-cell imaging (500 ms for 60 s) of HeLa cells co-expressing GFP-BIN1 (e) or GFP-BIN1 SH3-ΔC (f) together with mCherry-dynamin (see Methods). White arrows point to fission events observed in (e). Kymographs were obtained along the blue lines. Measurements were performed from at least three independent experiments. Box plots display the average value (solid square, also shown on top of each box plot), the median value (solid line), the s.d. and the interquartile range (25–75%) (n=39, n=27 and n=22, for control, N-BAR and N-BAR D151N, respectively). Error bars represent s.d. ***P<0.001 (t-test). Scale bars, 20 μm for flat membranes, 10 μm for cell imaging and 2 μm for kymographs.

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