Fig. 1. Adhesion ring localization at invadopodia correlates with invadopodium-associated extracellular matrix degradation.

(A) Confocal images of CA1d cells cultured on 1% crosslinked gelatin/FITC-FN (“Matrix”, green in triple merge) and immunostained for vinculin (Vinc) or paxillin (Pax) to mark adhesion structures (red in merges) and actin or cortactin (Cort) to mark invadopodia (blue in merges). Example focal adhesions (black arrows) and adhesion rings surrounding invadopodia (white arrows) are indicated. Dark areas in the matrix are where the FITC-FN/gelatin was degraded. Scale bars  =  5 µm. (B) Zooms of boxed areas in A. Scale bars  =  5 µm. (C) Percentage of adhesion ringed- and non-adhesion ringed-invadopodia colocalized with degradation of the underlying matrix in CA1d or SCC61 cells stained for paxillin and cortactin. Data are compiled from cells plated on soft PA, hard PA and glass conditions. Data separated by condition are shown in supplementary material Fig. S1. n≥180 cells per condition from 3 independent experiments. (D) Spearman correlation of the invadopodium ring or focal adhesion characteristics of an individual cell with the degradation area of FITC-FN underneath the same cell for CA1d or SCC61 cells plated on the noted underlying surfaces and stained for paxillin and cortactin. n≥60 cells per condition from 3 independent experiments. *P<0.05; **P<0.01; ***P<0.001 vs. no correlation.