LKB1 limits single-cell and collective cell migration in melanoma. (A) Single-cell speed of LKB498 cells in 1 mg/ml 3D collagen. (null, n = 73; addback, n = 83.) (B) Spheroid invasion of LKB498 cells into 3.2 mg/ml 3D collagen. Cells were stained with Alexa Fluor 568–phalloidin to label actin and Hoechst 33342 to label nuclei. (C) Spheroid outgrowth of LKB498 cells into increasing concentrations of 3D collagen. (1 mg/ml: null, n = 22; addback, n = 24. 2 mg/ml: null, n = 21; addback, n = 27. 4 mg/ml: null, n = 30; addback, n = 29. 6 mg/ml: null, n = 20; addback, n = 23. 8 mg/ml: null, n = 28; addback, n = 23.) (D) Single-cell speed of LKB498 cells on increasing fibronectin concentrations. (0.1 µg/ml: null, n = 94; addback, n = 104. 1 µg/ml: null, n = 99; addback, n = 96. 10 µg/ml: null, n = 149; addback, n = 162. 100 µg/ml: null, n = 134; addback, n = 150.) (E) Montage images and edge outlines of time-lapse scratch wounds. (F) Relative wound area over time. (LKB498: null, n = 43; addback, n = 32. TKL2: null, n = 33; addback, n = 33.) Gray shaded area indicates times at which relative wound area shows significant differences with P < 0.01. Insets show magnified plots of time points at which pausing occurs (represented by the bars) at the wound boundary. (G) Scratch wound of LKB498 LKB1 addback monolayer on Cy3-fibronectin (FN). DIC, differential interference contrast. All data are shown as means ± SEM. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001 by two-tailed unpaired t test. Bars, 100 µm.