Figure 3.
Figure 3. The Hippo pathway is required to polarize actin and promote migration. (A and B) Confocal micrograph of a control egg chamber at stage 9 (A) or 10 (B) labeled with phalloidin (red) to visualize the actin cytoskeleton, GFP (green) to mark the mutant clones of cells induced with the MARCM technique, and DAPI (blue) to stain all nuclei. Insets show F-actin staining of clusters at high magnification. (C) Stage 10 egg chamber with a cluster (arrow) composed entirely of exAP50 mutant border cells (FRT/FLP, GFP negative). Inset shows F-actin accumulating inside the cluster. (D) Stage 10 egg chamber with a cluster (arrow) composed entirely of exAP50 kib32 double-mutant border cells (FRT/FLP, GFP negative). Inset shows the cluster architecture completely fails to form. (E) Stage 10 migration index for quantitation of border cell migration. (F) Quantification of the stage 10 migration index for the following genotypes: control (n > 100), kib32 (n = 36), exAP50 (n = 34), and exAP50 kib32 (n = 8). (G and H) Stage 9 (G) and stage 10 (H) egg chambers with clusters (arrows) composed entirely of hpo42-47 mutant border cells (GFP positive). (I and J) Stage 9 (I) and stage 10 (J) egg chambers with clusters (arrows) composed entirely of wtsX1 mutant border cells (GFP positive). hpo42-47 and wtsX1 mutant border cell clusters both show delayed migration and F-actin polarization defects (insets). The hpo42-47 and hpoJM1 alleles display the same border cell migration delay phenotype. (K) Quantification of the stage 10 migration index of the following genotypes: control (n > 100), hpoJM1 (n = 94), and wtsX1 (n = 93). In each case, only clusters in which all border cells were mutant for a given allele (GFP positive) were analyzed; n, number of egg chambers examined. (L) Quantification of average F-actin and P-MyoII staining intensity levels at the outer rim versus inner membranes in control and wts mutant clusters (WT clusters, n = 5; wts clusters n = 12). p-MyoII staining in control and wts mutant clusters is also shown. Bars, 50 µm (5 µm for insets).

The Hippo pathway is required to polarize actin and promote migration. (A and B) Confocal micrograph of a control egg chamber at stage 9 (A) or 10 (B) labeled with phalloidin (red) to visualize the actin cytoskeleton, GFP (green) to mark the mutant clones of cells induced with the MARCM technique, and DAPI (blue) to stain all nuclei. Insets show F-actin staining of clusters at high magnification. (C) Stage 10 egg chamber with a cluster (arrow) composed entirely of exAP50 mutant border cells (FRT/FLP, GFP negative). Inset shows F-actin accumulating inside the cluster. (D) Stage 10 egg chamber with a cluster (arrow) composed entirely of exAP50 kib32 double-mutant border cells (FRT/FLP, GFP negative). Inset shows the cluster architecture completely fails to form. (E) Stage 10 migration index for quantitation of border cell migration. (F) Quantification of the stage 10 migration index for the following genotypes: control (n > 100), kib32 (n = 36), exAP50 (n = 34), and exAP50 kib32 (n = 8). (G and H) Stage 9 (G) and stage 10 (H) egg chambers with clusters (arrows) composed entirely of hpo42-47 mutant border cells (GFP positive). (I and J) Stage 9 (I) and stage 10 (J) egg chambers with clusters (arrows) composed entirely of wtsX1 mutant border cells (GFP positive). hpo42-47 and wtsX1 mutant border cell clusters both show delayed migration and F-actin polarization defects (insets). The hpo42-47 and hpoJM1 alleles display the same border cell migration delay phenotype. (K) Quantification of the stage 10 migration index of the following genotypes: control (n > 100), hpoJM1 (n = 94), and wtsX1 (n = 93). In each case, only clusters in which all border cells were mutant for a given allele (GFP positive) were analyzed; n, number of egg chambers examined. (L) Quantification of average F-actin and P-MyoII staining intensity levels at the outer rim versus inner membranes in control and wts mutant clusters (WT clusters, n = 5; wts clusters n = 12). p-MyoII staining in control and wts mutant clusters is also shown. Bars, 50 µm (5 µm for insets).

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