Fig. 7: CASC4 N-terminal domain (NTD) induces podosome-like structures.

a, b Immunofluorescence analysis and quantification of MDA-MB-231 stable cells stained for phalloidin (F-actin; white labeling) or tyrosine kinase substrate 5 (TKS5; green labeling). c Schematic representation of CASC4 truncated mutant (NTD) with a 3xflag in N-terminal and SP-ΔTM-CASC4 with a V5 in C-terminal. The blue circles are depicting potential N-glycosylation sites and the white circles are depicting O-glycosylation sites. d Western blot analysis of cell lysates and media from MDA-MB-231 cells transiently transfected with different CASC4 constructs. e Immunofluorescence analysis of transiently transfected MDA-MB-231 cells stained for phalloidin (F-actin; white labeling), tyrosine kinase substrate 5 TKS5 (green labeling), and nucleus stained with DAPI (blue labeling). Yellow arrows represent colocalization between F-actin and TKS5. The ability of transiently transfected MDA-MB-231 cells to migrate (6 h) (e, f) or invade (16 h) (g, h) was assessed by counting the number of cells stained with DAPI on the underside of a polycarbonate membrane under a phase contrast microscope (×20).These results are representative of at least three independent experiments. Error bars indicate averaged values ± standard error from the mean (SEM). P-values: *P ≤ 0.05, **P < 0.01, ***P ≤ 0.001 n.s. not significant (two-sided Student’s t-test). Scale: 10 µm.