Additional file 2: of In vivo screening reveals interactions between Drosophila Manf and genes involved in the mitochondria and the ubiquinone synthesis pathway

Screen for genetic interaction partners of DmManf. A tiff file. A) Scheme of the crosses used in the partial RNAi library screen. First, randomly selected UAS-x-RNAi lines were crossed to wing-specific driver line MS1096-GAL4 (1). Lines showing similar phenotype to UAS-DmManf-RNAi lines were selected to the next stage. Second, two GAL4 drivers, wing-specific MS1096-GAL4 and semi-ubiquitous 69B-GAL4, were used in both wild type and heterozygous DmManf Δ96 mutant background (2) – UAS-x-RNAi lines showing distinct phenotypes in wild type and heterozygous DmManf Δ96 mutant backgrounds were selected. Secondary stage was repeated for selected UAS-x-RNAi lines in order to ensure the observed interactions. At stage 3, 69B-GAL4 and MS1096-GAL4 drivers were used to express UAS-x-RNAi lines with or without UAS-DmManf overexpression construct (3). Based on stages 2 and 3, 21 UAS-x-RNAi lines were selected as candidates for final stage 4. (4) UAS-x-RNAi lines were expressed with tub-GAL4 with and without DmManf overexpression to study whether high levels of DmManf affected ubiquitous silencing of selected genes (see Additional file 4). B-C) DmManf (magenta) was ubiquitously expressed in the wing disc of 3rd instar larvae. MS1096-GAL4 expression detected by UAS-mCD8-GFP (green) was found mainly in the dorsal wing compartment but also in other regions of the wing disc (B). MS1096-GAL4 expression pattern was also detected in the CNS (C). Nuclear counterstain DAPI (gray) was used to mark the tissue morphology. Scale bar 50 μm. D) Insertion of GAL4 construct GawB in driver lines 69B-GAL4 and MS1096-GAL4 [31] did not affect adult fly phenotype (+/+, top row). Heterozygous DmManf mutation (DmManf Δ96 /+, middle row) or overexpression of DmManf (UAS-DmManf L5 , bottom row) together with 69B-GAL4 and MS1096-GAL4 insertions showed no obvious phenotype in adult flies. (TIFF 3486 kb)