One-generation in vivo Drosophila genetic screening approach relevant to rhabdomyosarcoma

Galindo KA, Endicott TR, Avirneni-Vadlamudi U, Galindo RL. A rapid one-generation genetic screen in a Drosophila model to capture rhabdomyosarcoma effectors and therapeutic targets. G3 (Bethesda). 2014 Dec 9;5(2):205-17. PMID: 25491943; PMCID: PMC4321029.

From the abstract: "Rhabdomyosarcoma (RMS) is an aggressive childhood malignancy of neoplastic muscle-lineage precursors that fail to terminally differentiate into syncytial muscle. The most aggressive form of RMS, alveolar-RMS, is driven by misexpression of the PAX-FOXO1 oncoprotein ... Here, we report a new approach to dissect RMS, exploiting a highly efficient Drosophila PAX7-FOXO1 model uniquely configured to uncover PAX-FOXO1 RMS genetic effectors in only one generation. ... These studies show the utility of the PAX-FOXO1 Drosophila system as a robust one-generation (F1) RMS gene discovery platform and demonstrate how Drosophila transgenic conditional expression models can be configured for the rapid dissection of human disease. "

RNA-based mitochondrial targeting technique shown to work in vivo in flies and in human cells

Towheed A, Markantone DM, Crain AT, Celotto AM, Palladino MJ. Small mitochondrial-targeted RNAs modulate endogenous mitochondrial protein expression in vivo. Neurobiol Dis. 2014 Sep;69:15-22. PMID: 24807207; PMCID: PMC4106415.

From the abstract: “Endogenous mitochondrial genes encode critical oxidative phosphorylation components and their mutation results in a set of disorders known collectively as mitochondrial encephalomyopathies. There is intensive interest in modulating mitochondrial function as organelle dysfunction has been associated with numerous disease states. Proteins encoded by the mitochondrial genome cannot be genetically manipulated by current techniques. Here we report the development of a mitochondrial-targeted RNA expression system (mtTRES) utilizing distinct non-coding leader sequences (NCLs) and enabling in vivo expression [in Drosophila] of small mitochondrial-targeted RNAs. mtTRES expressing small chimeric antisense RNAs was used as translational inhibitors (TLIs) ... This technique has important and obvious research and clinical implications.”

Small ORF-related genome-wide screen performed at the DRSC reported by Zanet, Benrabah et al. in Science

Zanet J, Benrabah E, Li T, Pélissier-Monier A, Chanut-Delalande H, Ronsin B, Bellen HJ, Payre F, Plaza S. Pri sORF peptides induce selective proteasome-mediated protein processing. Science. 2015 Sep 18;349(6254):1356-8. PMID: 26383956.

From the abstract: "A wide variety of RNAs encode small open-reading-frame (smORF/sORF) peptides, but their functions are largely unknown. Here, we show that Drosophila polished-rice (pri) sORF peptides trigger proteasome-mediated protein processing, converting the Shavenbaby (Svb) transcription repressor into a shorter activator. A genome-wide RNA interference screen identifies an E2-E3 ubiquitin-conjugating complex, UbcD6-Ubr3, which targets Svb to the proteasome in a pri-dependent manner. ... "

S2 cells used to express human HER2 protein domains

Kanthala S, Mill CP, Riese DJ 2nd, Jaiswal M, Jois S. Expression And Purification Of HER2 Extracellular Domain Proteins In Schneider2 Insect Cells. Protein Expr Purif. 2015 Sep 9. PMID: 26363121.

CRISPR + RNAi screening in Drosophila cells points to potential new drug targets for treatment of tuberous sclerosis complex (TSC)

Housden BE, Valvezan AJ, Kelley C, Sopko R, Hu Y, Roesel C, Lin S, Buckner M, Tao R, Yilmazel B, Mohr SE, Manning BD, Perrimon N. Identification of potential drug targets for tuberous sclerosis complex by synthetic screens combining CRISPR-based knockouts with RNAi. Sci Signal. 2015 Sep 8;8(393):rs9. PMID: 26350902.

From the abstract: "The tuberous sclerosis complex (TSC) family of tumor suppressors, TSC1 and TSC2, function together in an evolutionarily conserved protein complex ... Mutation or aberrant inhibition of the TSC complex is common in various human tumor syndromes and cancers. The discovery of novel therapeutic strategies to selectively target cells with functional loss of this complex is therefore of clinical relevance ... We developed a CRISPR-based method to generate homogeneous mutant Drosophila cell lines. By combining TSC1 or TSC2 mutant cell lines with RNAi screens against all kinases and phosphatases, we identified synthetic interactions with TSC1 and TSC2. Individual knockdown of three candidate genes ... reduced the population growth rate of Drosophila cells lacking either TSC1 or TSC2 but not that of wild-type cells ... [and] had similar growth-inhibiting effects in mammalian TSC2-deficient cell lines, including human tumor-derived cells, illustrating the power of this cross-species screening strategy to identify potential drug targets."