Figure 3. Unrestricted Upd-exposure induces defects in both normal and Lgl-KD follicle cells.

(A) Confocal images of stage-8 egg chambers containing RFP⁺ (red) follicle cells with Lgl-KD+N^ICD (Left) and Lgl-KD+Upd-OE (Right) expression. Eya expression is detected within immature cells (green). Oocyte are marked by asterisks (*). Intensities of RFP expression and Eya antibody enrichment are shown separately (grayscale) in the middle and bottom panels. (B) Box-and-whisker plot to show the distribution of ovarioles from different genotypes (X-axis) with complete or partial (<2 layers) rescue of multilayering over 3 independent experiments. (C) Confocal images of egg chambers containing follicle cells expressing tj^TS-driven RFP control (1^st row), Upd-OE for 72h (2^nd row), and Lgl-KD+Upd-OE for 72h (3^rd row) and 24h (4^th row) during early and midoogenesis. Abnormal stalk cells are indicated by yellow arrows while fused egg chambers with encapsulation errors are indicated by yellow arrowheads. Nuclei is marked by DAPI (blue). F-Actin is marked by Phalloidin (white). Scale bars: 20μm. (D) Box-and-whisker plot to show the number of egg chambers containing immature (Eya+) follicle cells (Y-axis) in individual ovariole, for when follicle cells express Upd-OE (compared with control). The p-value obtained from unpaired t-test comparison of the two distributions is indicated above the plot. (E) Bar plot to show the occurrence (%) of stage-specific ovarian phenotypes when Upd-OE and Lgl-KD+Upd-OE are expressed in follicle cells for 24h and 96h (compared with ovarioles with tj^TS control follicle cells at 96h). (F) PCA distribution of whole-tissue transcriptomes of ovaries containing follicle cells expressing tj^TS-driven Lgl-KD, Upd-OE and Lgl-KD+Upd-OE for short (24h) and long (96h) time-points, along with the corresponding control samples. Individual samples are uniquely colored and replicates per sample are encircled. See also Figure S5.