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. Author manuscript; available in PMC: 2023 Dec 22.
Published in final edited form as: Cell Rep. 2023 Oct 19;42(10):113302. doi: 10.1016/j.celrep.2023.113302

Figure 6. FAP knockdown blocks invadopodia precursor stabilization and subsequent maturation.

Figure 6.

(A, D, E, H, F, G) SCC61 cells expressing the indicated FAP shRNAs were seeded onto gelatin-coated coverslips for 48 h, and stained for F-actin (magenta), and cortactin (green) (A), Tks5 (green) (E), MT1-MMP (green) (H). N = >20 cells per group. Scale bar, 10 μm. Quantification of normalized fluorescence intensity of cortactin (D), Tks5 (F), and MT1-MMP (G) at invadopodia relative to control shRNA.

(B) Quantification of invadopodia protrusion count per cell in SCC61 cells expressing FAP shRNAs.

(C) Quantification of F-actin intensity in SCC61 cells expressing different FAP shRNAs.

(I) Quantification of average invadopodia protrusion displacement divided by its lifetime (number of frames) (pixels/min) in Life-Act mCherry expressing SCC61 cells with FAP shRNAs. Each dot in the graph represents individual precursors. N > 100 precursors per group.

(J) Polar graph indicating invadopodia precursor trajectories in a representative SCC61 cell expressing shRNAs targeting FAP. The outer ring (green) represents the distance of 4 μm and inner circle (blue) represents distance of 2 μm. Tracks were measured at 20 s/frame for 30 min.

(K) Time lapse pictures of live SCC61 cells expressing F-actin and FAP shRNAs captured using TIRF microscopy at 100× magnification at time 0 (green) and at 1 h (magenta). Insets show zoomed in view of invadopodia displacement over time. Scale bar, 10 μm. Error bars are the SEM. ***p < .001, **p < .01, *p < .05. ns, not significant (p > 0.5). Student’s t-test. See also Figure S4A and Videos S1−S3.