Results: 5

1.
Figure 2

Figure 2. From: Light-mediated activation reveals a key role for Rac in collective guidance of cell movement in vivo.

Forward or backward movement in response to photoactivatable Rac.
(a-i) In an otherwise wild-type background, PA-RacQ61L can promote forward (a-c) or backward (d-i) movement. (j-l) Forward and (m-o) reverse migration of border cells expressing PVRDN, EGFRDN, and PA-RacQ61L. The schematics at the right show the position of the cluster within the egg chamber. Scale bars, 20μ. In panels with two colors, red represents the starting position and green shows the ending position over the indicated time period. p) Average migration speeds for clusters expressing the indicated proteins in response to illumination of the front or the back of the cluster. PA-Rac refers to PA-RacQ61L. C450M is the light-insensitive control. Values represent the average of the indicated number (N) of experiments and error bars show the standard deviation.

Xiaobo Wang, et al. Nat Cell Biol. ;12(6):591-597.
2.
Figure 4

Figure 4. From: Light-mediated activation reveals a key role for Rac in collective guidance of cell movement in vivo.

Local photoactivation or photoinactivation of Rac in one cell affects the morphology and behavior of other cells in the group.
(a-l) Confocal images of border cell clusters before (0 min) and after (60 min) photoactivation. Circles indicate areas of laser treatment. The white arrow in panel l indicates the direction the border cells would normally migrate and applies to all panels. Scale bar is 10 μm. In c, f, i and l, red shows the starting position and green shows the ending position. m, The average number of cells sending protrusions simultaneously within one cluster was calculated from 3-D reconstructed images (see methods and Figure S6). “−” and “+” indicate before and after photoactivation. n, Directionality indices were calculated from the same samples (see methods).

Xiaobo Wang, et al. Nat Cell Biol. ;12(6):591-597.
3.
Figure 3

Figure 3. From: Light-mediated activation reveals a key role for Rac in collective guidance of cell movement in vivo.

Responsiveness of border cells to PA-RacQ61L depends on their location within the egg chamber.
(a-c) Within the anterior third of the egg chamber, photoactivation diverts border cells. (d-i) In the middle third, photoactivation has little effect. (c) In the posterior third, photoactivation again drives border cells toward the side. Scale bars represent 20 μm, elapsed time is shown in minutes. Schematics show border cell position within the egg chamber. In panels with two colors, red indicates the starting position and green shows the ending position. (m) Summary of experiments. The lengths of the arrows indicate the average distance migrated in the indicated direction in response to PA-RacQ61L, for border cells starting at the base of the arrow. The black arrow indicates the normal migration direction. Yellow Xs indicate positions beyond which border cells did not move. Each arrow summarizes at least five experiments.

Xiaobo Wang, et al. Nat Cell Biol. ;12(6):591-597.
4.
Figure 5

Figure 5. From: Light-mediated activation reveals a key role for Rac in collective guidance of cell movement in vivo.

Rac activity pattern in migrating border cell clusters.
(a-e’) A time lapse series of migrating border cells. (a-e) YFP channel only (a’-e’) processed FRET signal f. FRET image of wild-type border cells displayed with Red-Hot pseudocolor divided into 30 sectors. The yellow line shows the direction of migration. The white circle indcates the central region that was excluded from the analysis. (g) Average FRET efficiency from image f plotted in a radar map. Efficiency higher than 1.2 is highlighted in purple. (h-j) Representative FRET patterns in wild-type (h) and EGFRDN- and PVRDN-expressing (j) border cells. (i, k) Heatmaps from 30 examples of each genotype. Each row represents the FRET signal distribution of an individual border cell cluster. Positions from −15 to 15 plotted on the x-axis correspond to the sectors, where 0 represents the front of the cluster. l. FRET efficiencies in border cells of the indicated genotypes. All results were normalized to the efficiency of RacDN. (m) Distributions of average FRET efficiencies in wild-type (blue) and PVRDN/EGFRDN border cells, plotted as a function of sector number, where 0 represents the front.

Xiaobo Wang, et al. Nat Cell Biol. ;12(6):591-597.
5.
Figure 1

Figure 1. From: Light-mediated activation reveals a key role for Rac in collective guidance of cell movement in vivo.

Local activation of PA-Rac1 redirects an entire border cell group.
(a-c) Egg chambers labelled with DAPI (blue) to stain all nuclei, Alexa 488-phalloidin (green) to mark actin filaments, and mCherry (red) to show PA-RacQ61L. (d-f), Higher magnification views of border cells from each stage. (g-i) PA-RacQ61L expression only. (j) Schematic diagram from8 showing the mechanism of PA-Rac light-activation. (k) Schematic of border cell cluster composed of two non-migratory polar cells (purple, p) which do not express slbo-Gal4 and are therefore unlabeled in all subsequent images. Polar cells are surrounded by 4-6 migratory border cells (green, b). (l-t) Selected still images from a time-lapse film of the response of border cells to photoactivation of PA-RacQ61L. (l-n) Photoactivation diverts border cells to the edge of the egg chamber. (o-q) Continued photoactivation in same direction did not move them further along the edge. (r-t) Photoactivation of the same cluster in a different position drove movement towards the egg chamber center. In n, q and t the starting position of the cluster is shown in red and the final position in green. Schematics at right show the position of the treated cluster within the egg chamber. Red boxes indicate the regions shown in the micrographs. Red arrow indicates the normal direction of migration. Pink arrow shows the direction the cells move if they respond to the light. (u-w) Phototreatment of light insensitive control C450M-PA-Rac1Q61L. In l, n, q and u, solid arrows indicate the normal direction of migration; circles indicate where the laser light was applied. Dashed arrows indicate the direction the cells move if they respond to the light. Scale bars, 20 μm.

Xiaobo Wang, et al. Nat Cell Biol. ;12(6):591-597.

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