|
Fig. 1 SERCA inhibition disrupts Drosophila air sac via cell-autonomous defects in epithelial migration and proliferation. (A) serca RNAi disrupts airway morphogenesis. Wild-type air sac (left), with GFP-labeled respiratory epithelium. serca RNAi, driven by breathless in respiratory epithelium, results in a stunted (right) or absent air sac. Scale bar: 25 μm. (Also see Fig. S2.) (B) serca RNAi alters the position of cells expressing distal marker, escargot (red), which normally are positioned only at the air sac tip (top panels). In serca RNAi mutants (bottom), escargot-expressing cells are retained also in the malformed air sac stalk (arrows, right). breathless-dependent GFP expression identifies the wild-type air sac (top row, left) and abnormal serca RNAi epithelial air sac (bottom, left). Scale bar: 20 μm. (C) serca-deficient cells are necessary within the air sac to disrupt budding. FLP-recombinase to generate serca RNAi clones (green) shows that if no such clones are induced in the air sac it forms normally (top, epithelium in red), whereas when flipped cells populate the air sac, it fails to develop properly (bottom). Scale bar: 25 μm. (D) Flip-out serca RNAi cells have a cell-autonomous migration defect. Plot shows proportions of flipped cells localized to the tip, middle and stalk thirds of the air sac for control and serca RNAi clones. serca RNAi cells are relatively excluded from the air sac tip and restricted more to the stalk. (E) Flip-out serca RNAi cells have a cell-autonomous proliferation defect. Plot of number of cells per contiguous group of flipped cells for control and serca RNAi clones. Median (interquartile range) of phosphohistone H3 (PH3)-positive cells per air sac: 2.5 (1-4) control vs 0 (0-1) serca RNAi (n=18 per group; P<0.001, Mann–Whitney).