Figure 5. Effects of local activation of optoPlexin on cell morphology and motility.

Effects on the illuminated protrusions (a), measured in a 50 μm-diameter circle centred at the region of illumination, and total cell areas (b) after 7.5 min of local illumination in MC3T3-E1 cells expressing Cry2, optoPlexin, optoPlexin-delPBD, optoPlexin and pretreated with 10 μM Y-27632 ROCK inhibitor, and optoPlexin-RA, n=9–14 cells. (c) Cell centroid velocities at different times upon local activation of optoPlexin in MC3T3-E1 cells. Blue line, illumination at 440 nm, n=21 cells. (d) The locally induced retraction (blue) and the distal protrusion (red) after pulses of illumination (yellow) of an optoPlexin-expressing cell was illustrated in a morphology diagram. The overlapping cell area before and after illumination was labeled in green. Based on the centroids of these colour-coded regions, θ₁ and θ₂ were used to describe the angles of retraction and protrusion relative to the direction from the centroid of the cell at time 0 to the centre of illumination, respectively. Similarly, d₁ and d₂ indicated their centroid distances to the region of illumination, respectively. (e,f) Angles of retraction and protrusion induced by local activation of optoPlexin in MC3T3-E1 cells and (g) s.d. of their distribution, n=14 cells. (h) Proximity of retractions and protrusions induced by local activation of optoPlexin in MC3T3-E1 cells to the region of illumination, n=14 cells. For a,b,c,h, means±s.e.m are shown, ***P<0.001 and *P<0.05, NS, not significant, Student’s t-test. For g, ***P<0.001 and *P<0.05, NS, not significant, F-test.