Fig. 5

Scheme of the quantitative migration assay. a Cells are seeded on 35 µm round patterns. After complete adhesion, the adhesive reagent BCN-RGD is added and binds to the coverslip’s surface, allowing free 2D cell migration (top). Directed migration can be triggered by optogenetic activation of GEFs through light gradients at the same time as cell adhesion is released (bottom). b Examples of cells expressing CIBN-GFP-CAAX and TIAM-CRY2-mCherry with (3× gradient, bottom) or without (top) photo-activation (visualized: TIAM-CRY2-mCherry). Time indicates the duration after addition of BCN-RGD and concomitant blue light illumination. The dashed orange line corresponds to the initial position of the cell center. c, d HeLa cells expressing CIBN-GFP-CAAX and ITSN-CRY2-mCherry or CIBN-GFP-CAAX and TIAM-CRY2-mCherry were illuminated with various gradients of light as the adhesive patterns were released. c Average morphodynamic maps for each condition (ITSN: top, TIAM: bottom). The vertical axis corresponds to the coordinate along the cell contour (centered on the direction of the light gradient) and the horizontal axis corresponds to time. The local velocity of the edge of the cell membrane is color coded accordingly to the bar on the right side. Gradient extents are schemed on the left side of each map. ITSN: n=25 (control with uniform illumination), n=16 (1× gradient), n=20 (2×), n = 19 (3×) or n = 16 (4×). TIAM: n = 19 (ctrl), n = 18 (1×), n = 18 (2×), n = 17 (3×), n = 18 (4×). d We tracked the position of the centroid of individual cells. Top: Trajectories of cells stimulated with various gradients of ITSN. Bottom: The angles between the displacement vector (initial to final centroid position) and the stimulation axis for each cell are represented in polar coordinates. Scale bars: 20 µm