Fig. 10: NHSL1 reduces the stability of lamellipodia protrusion. | Nature Communications

Fig. 10: NHSL1 reduces the stability of lamellipodia protrusion.

From: Nance-Horan Syndrome-like 1 protein negatively regulates Scar/WAVE-Arp2/3 activity and inhibits lamellipodia stability and cell migration

Fig. 10

a, b Movie stills showing protrusion (blue) and retraction (red) dynamics of wild-type B16-F1, NHSL1 CRISPR 2 and B16-F1 cells overexpressing Myc-tagged NHSL1. Scale bar: 20 μm. c, d NHSL1 reduces the length of the longest uninterrupted lamellipodium. Each data point represents the mean length of the longest uninterrupted lamellipodium quantified from all frames of a movie of one LifeAct-EGFP expressing B16-F1 cell (5 s intervals, 10 min duration) (see ‘Methods’ for details). Control (black circles), NHSL1 CRISPR 2 (magenta squares), 21 (green triangles) (c) and control (black circles) and Myc-NHSL1 overexpression (red crosses) (d). c Control: n = 13; NHSL1 CRISPR 2: n = 12; NHSL1 CRISPR 21: n = 13 cells. One-way ANOVA: P = 0.0111; F(2,35) = 5.133; Dunnett’s test *P = 0.0144, control versus CRISPR 21: not significant: nsP = 0.9949. d control n =10; Myc-NHSL1 n = 9 cells. ***P = 0.006, Mann–Whitney test. e, f Quantification (see ‘Methods’ section) of lamellipodia protrusion speed (e) and lamellipodia stability (f) of randomly migrating wild-type B16-F1 cells or CRISPR 2 cells expressing either NHSL1 (NHSL1 WT) or the NHSL1 mutant in the Scar/WAVE complex binding sites (NHSL1 SW Mut) or Myc alone (control) plated on laminin. WT Control: n = 13, NHSL1 CRISPR 2: n = 12, Rescue Myc-NHSL1 SW Mut: n = 11, Rescue Myc-NHSL1 SW WT: n = 15 cells. One-way ANOVA: P = 0.2097, F(3,47) = 1.568; Dunnett’s test: NHSL1 CRISPR 2 vs Rescue SW mut: nsP = 0.2043; NHSL1 CRISPR 2 vs Rescue SW WT: nsP = 0.1573; NHSL1 CRISPR 2 vs WT control: nsP =0.7639. f The standard deviation of the cone speed shows the fluctuation of speeds along the edge and serves as a measure for the stability of lamellipodial protrusions. WT control: n = 13, NHSL1 CRISPR 2: n = 12, Rescue Myc-NHSL1 SW Mut: n = 11, Rescue Myc-NHSL1 SW WT: n = 15 cells. One-way ANOVA: P = 0.0412, F(3,47) = 2.971; Dunnett’s test: NHSL1 CRISPR 2 vs Rescue SW mut: nsP = 0.8865; NHSL1 CRISPR 2 vs Rescue SW WT: *P = 0.0299; NHSL1 CRISPR 2 vs WT control: nsP = 0.1173. c–f Results are mean ± SEM (error bars), four independent biological repeats. Source data are provided as a Source Data file.

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