Fig. 7

Junction dynamics upon VEGF treatment in confluent HUVECs. HUVECs expressing (a) VE-cadherin-EGFP (b–e) LifeAct-EGFP or (f–g) EGFP-p20, respectively, were treated with 50 ng ml⁻¹ VEGF. a (upper) Controls display regular VE-cadherin-EGFP dynamics with VE-cadherin plaques (white arrows) due to JAIL formation. (middle) After VEGF treatment, elongated cells exhibit a polar distribution with an interrupted VE-cadherin pattern at cell poles (red arrowheads) followed by transient formation of new large VE-cadherin plaques (dotted lines). (lower) Lateral junctions display linear VE-cadherin pattern with appearance of small VE-cadherin plaques (white arrows). Small VE-cadherin interruptions (red arrows) appear transiently. Time scale: mm:ss; scale bars indicate 10 and 5 µm in the overview and cropped images. b LifeAct-EGFP dynamics after VEGF. (upper) The overview images demonstrate both polygonal cells and elongated cells (white arrows) (scale bar: 30 µm). (Area 1) Polygonal cells display constitutive junction remodelling that includes the transient appearance of small stress fibres (red arrows) and small JAIL (white arrowheads) without significant cell elongation. (Area 2) A progressively moving elongated cell exhibits many filopodia (empty arrowheads) that appear on the sides of large JAIL (white arrowheads). Note: stress fibres (red arrows) preferentially emerge at the leading edge of the cells. Time scale: hh:mm; scale bar: 10 µm. c, d JAIL and filopodia number in polygonal and elongated cells in VEGF-treated culture. e Track plots illustrate migration path of elongated cells (black line) and polygonal cells (red line). f Small JAIL formation at the lateral junctions (cropped area 1, arrowheads) and large JAIL development at the cell poles (cropped area 2, arrowheads) in VEGF-induced elongated ECs expressing EGFP-p20. The dotted line indicates the cell border; arrow indicates the direction of cell movement. Time scale, mm:ss; scale bar represent 20 and 10 µm in overview and cropped images. g JAIL size at the cell poles and the lateral sides of elongated cells induced by VEGF. 157 JAIL at the cell pole and 152 JAIL at the lateral junctions from three independent experiments were analysed over a period of time (30 min). Unpaired student’s t test was used for statistics analysis and error bars indicate ± SEM