Figure 4
Dynamics of podoplanin during invadopodia assembly and matrix degradation. (a) Podoplanin rings form following actin polymerisation. Live-cell imaging of HN5 cells expressing PDPN-GFP and Lifeact-Ruby cultured on 0.5% glutaraldehyde-crosslinked gelatin. Pictures represent zoomed areas of invadopodia dynamics over time. (b) Panels 1, 2, 3 are kymographs showing the lifetime of representative invadopodia from time-lapse imaging in a (boxes). White arrowheads indicate invadopodium formation time, and black arrowheads indicate the time of podoplanin ring assembly. (c) Podoplanin ring assembly precedes matrix degradation. Time-lapse confocal microscopy of HN5 cells expressing PDPN-GFP cultured on 0.5% glutaraldehyde-crosslinked TRITC-gelatin. (d) Panels 1 and 2 show Kymographs representing a subset of invadopodia from time-lapse in c (boxes). White arrowheads indicate the time of podoplanin ring assembly, whereas black arrowheads indicate the time of gelatin degradation appearance. (e) Graph represents the percentage of podoplanin-ringed invadopodia in the different populations of invadopodia. Invadopodia were classified into two main populations: precursors (newly formed invadopodia with no ability to degrade the matrix) and active (invadopodia actively degrading the matrix). Results shown are the means±s.e.m. of n⩾70 invadopodia (five cells) for each condition over two independent experiments. ***P<0.0001. Animate sequences of these data are shown in Supplementary Movies S1 and S2. Bars=5 μm.
