An RNAi Screen for Genes Required for Growth ofDrosophilaWing Tissue

Abstract

Cell division and tissue growth must be coordinated with development. Defects in these processes are the basis for a number of diseases, including developmental malformations and cancer. We have conducted an unbiased RNAi screen for genes that are required for growth in the Drosophila wing, using GAL4-inducible short hairpin RNA (shRNA) fly strains made by the Drosophila RNAi Screening Center. shRNA expression down the center of the larval wing disc using dpp-GAL4, and the central region of the adult wing was then scored for tissue growth and wing hair morphology. Out of 4,753 shRNA crosses that survived to adulthood, 18 had impaired wing growth. FlyBase and the new Alliance of Genome Resources knowledgebases were used to determine the known or predicted functions of these genes and the association of their human orthologs with disease. The function of eight of the genes identified has not been previously defined in Drosophila. The genes identified included those with known or predicted functions in cell cycle, chromosome segregation, morphogenesis, metabolism, steroid processing, transcription, and translation. All but one of the genes are similar to those in humans, and many are associated with disease. Knockdown of lin-52, a subunit of the Myb-MuvB transcription factor, or βNACtes6, a gene involved in protein folding and trafficking, resulted in a switch from cell proliferation to an endoreplication growth program through which wing tissue grew by an increase in cell size (hypertrophy). It is anticipated that further analysis of the genes that we have identified will reveal new mechanisms that regulate tissue growth during development.

Gcm alleviates the inflammatory phenotype induced by Toll activation inDrosophilahemocytes

“…At last, to assess the impact of cholinergic transmission on hemocytes, we monitored the number of larval hemocytes after inhibiting the expression of the choline-acetyltransferase ChAT in neurons. To inhibit ChAT, we used a UAS-ChAT-RNAi (53) and a ChAT-Gal4 to drive ChAT-RNAi in the cholinergic neurons (54), which led to a strong increase in the number of hemocytes ( Figure 3J ), indicating that cholinergic neurons modulate hemocyte homeostasis through secretion of acetylcholine. As controls, we used OK6-Gal4 for type 1 motoneurons (55), which mostly secrete glutamate (56, 57), 21-7-Gal4 and 109(2) 80-Gal4 drivers for multidendritic neurons, which are cholinergic (54) and regulate hemocyte localisation and proliferation at the dorsal stripes through activin signaling (58).…”

DrosophilapVALIUM10 TRiP RNAi lines cause undesired silencing of Gateway-based transgenes

“…In vivo RNAi-based genetic screens have proved very successful in identifying novel genes involved in developmental and pathologic processes in a variety of Drosophila somatic tissues when expressed using tissue-specific Gal4 driver lines (Graca et al , 2021; Lesch et al , 2010; Neely et al , 2010; Pletcher et al , 2019; Port et al , 2011; Rotelli et al , 2019; Rylee et al , 2022; Saj et al , 2010; Zeng et al , 2015; Zhou et al , 2019). Hence, we embarked on a candidate genetic screen aimed at identifying factors controlling genome stability.…”

DrosophilapVALIUM10 TRiP RNAi lines cause undesired silencing of Gateway-based transgenes

“…Unexpected, off-target silencing of the RNase H1::GFP reporter by multiple pVALIUM10 RNAi lines In vivo RNAi-based genetic screens have proved very successful in identifying novel genes involved in developmental and pathologic processes in a variety of Drosophila somatic tissues when expressed using tissue-specific Gal4 driver lines (Lesch et al, 2010;Neely et al, 2010;Saj et al, 2010;Port et al, 2011;Zeng et al, 2015;Pletcher et al, 2019;Rotelli et al, 2019;Zhou et al, 2019;Graca et al, 2021;Rylee et al, 2022). Hence, we embarked on a candidate genetic screen aimed at identifying factors controlling genome stability.…”