The effects of the net advancement speed of the cell edge on the contact time between the lamellipodium and a filopodia adhesion.

A. Scatter plot of the part of the contact time of a filopodia adhesion with the advancing lamellipodium, with respect to the net advancement speed of the cell edge. All data were for REF52-β3-integrin-EGFP cells spreading on FN coated glass. Black: maturing filopodia adhesions. Red: disassembling filopodia adhesions. Due to the fast net advancement speed of the cell edge, the disassembling filopodia adhesions demonstrated a reduced contact time with the advancing lamellipodia, which subsequently had very brief of no pausing at these filopodia adhesions. Open symbols correspond to the filopodia adhesions in B–b. B. a. A fast spreading REF52 fibroblast expressing β3-integrin-EGFP (green) on FN coated glass (5 min–9 min 30 s after plating, Video S8). The cell membrane was stained with the fluorophore DiI (red). b. Magnified views of the dashed rectangle regions (I and II) in a. The selected frames from the time-lapse sequences showed the disassembly (I, horizontal arrows) or steady maturation (II, vertical arrows) of the filopodia adhesions as a result of ongoing advancement or prolonged pausing of lamellipodium at the two filopodia adhesions respectively. c. The advancement of cell two edges are represented by the kymographs that were generated along the corresponding kymograph lines (colored arrows in a) from the membrane fluorescence signal at the two filopodia adhesions. The dashed lines indicate the advancing (yellow) and pausing (pink) phases of lamellipodia at the respective filopodia adhesions. The net advancement speeds of the cell edges were measured at the corresponding sections of the kymographs as indicated by the dashed yellow lines (B-c I, 161 nm/s; B–c II, 147 nm/s). Scale bars: 2 µm (B–b), 5 µm (B–a).