Monday, June 4, 2018

in vivo RNAi screen focused on the female germline

Cho Y, Lai CM, Lin KY, Hsu HJ. A Targeted RNAi Screen Reveals Drosophila Female-Sterile Genes That Control the Size of Germline Stem Cell Niche During Development. G3 (Bethesda). 2018 May 15. PMID: 29764959.

From the abstract: "... we conducted a small-scale RNAi screen of 560 individually expressed UAS-RNAi lines with targets implicated in female fertility. RNAi was expressed in the soma of larval gonads, and screening for reduced egg production and abnormal ovarian morphology was performed in adults. Twenty candidates that affect ovarian development were identified and subsequently knocked down in the soma only during niche formation. ..."

Genome-wide cell-based RNAi screen related to JAK/STAT signaling

Fisher KH, Fragiadaki M, Pugazhendhi D, Bausek N, Arredondo MA, Thomas SJ, Brown S, Zeidler MP. A genome-wide RNAi screen identifies MASK as a positive regulator of cytokine receptor stability. J Cell Sci. 2018 May 30. PMID: 29848658.

From the abstract: "... In order to transduce ligand activation, cytokine receptors must dimerise. However, mechanisms regulating their dimerisation are poorly understood. In order to better understand the processes regulating cytokine receptor levels, activity and dimerisation, we used the highly conserved JAK/STAT pathway in Drosophila, which acts via a single receptor, known as Domeless. We have performed a genome-wide RNAi screen in Drosophila cells, identifying MASK as a positive regulator of Domeless dimerisation and protein levels. ... our results identify MASK as a novel regulator of cytokine receptor levels, and suggest functional conservation, which may have implications for human health."

Thursday, May 31, 2018

New in vivo RNAi screen report -- larval chordotonal organ morphogenesis

An RNAi Screen Identifies New Genes Required for Normal Morphogenesis of Larval Chordotonal Organs

Abeer Hassan, Yael Timerman, Rana Hamdan, Nitzan Sela,  Adel Avetisyan,  Naomi Halachmi and Adi Salzberg
G3: GENES, GENOMES, GENETICS June 1, 2018 vol. 8 no. 6 1871-1884

From the abstract: "... Here we describe an RNAi screen that focused on the ChO’s accessory and attachment cells and was performed in 2nd instar larvae to allow for phenotypic analysis of ChOs that had already experienced mechanical stresses during larval growth. Nearly one thousand strains carrying RNAi constructs targeting more than 500 candidate genes were screened for their effects on ChO morphogenesis. The screen identified 31 candidate genes whose knockdown within the ChO lineage disrupted various aspects of cell fate determination, cell differentiation, cellular morphogenesis and cell-cell attachment. ..."

in vivo fly RNAi screen identifies ortholog of SPO11 DNA break protein as pro-tumor in a fly tumor model

Rossi F, Molnar C, Hashiyama K, Heinen JP, Pampalona J, Llamazares S, Reina J, Hashiyama T, Rai M, Pollarolo G, Fernández-Hernández I, Gonzalez C. An in vivo genetic screen in Drosophila identifies the orthologue of human cancer/testis gene SPO11 among a network of targets to inhibit lethal(3)malignant brain tumour growth. Open Biol. 2017 Aug;7(8). pii: 170156. PMID: 28855394; PMCID: PMC5577452.

The abstract: "Using transgenic RNAi technology, we have screened over 4000 genes to identify targets to inhibit malignant growth caused by the loss of function of lethal(3)malignant brain tumour in Drosophila in vivo. We have identified 131 targets, which belong to a wide range of gene ontologies. Most of these target genes are not significantly overexpressed in mbt tumours hence showing that, rather counterintuitively, tumour-linked overexpression is not a good predictor of functional requirement. Moreover, we have found that most of the genes upregulated in mbt tumours remain overexpressed in tumour-suppressed double-mutant conditions, hence revealing that most of the tumour transcriptome signature is not necessarily correlated with malignant growth. One of the identified target genes is meiotic W68 (mei-W68), the Drosophila orthologue of the human cancer/testis gene Sporulation-specific protein 11 (SPO11), the enzyme that catalyses the formation of meiotic double-strand breaks. We show that Drosophila mei-W68/SPO11 drives oncogenesis by causing DNA damage in a somatic tissue, hence providing the first instance in which a SPO11 orthologue is unequivocally shown to have a pro-tumoural role. Altogether, the results from this screen point to the possibility of investigating the function of human cancer relevant genes in a tractable experimental model organism like Drosophila."

Tuesday, May 8, 2018

Drosophila cell-based RNAi screen related to treatment of epilepsy

Lin WH, He M, Fan YN, Baines RA. An RNAi-mediated screen identifies novel targets for next-generation antiepileptic drugs based on increased expression of the homeostatic regulator pumilio. J Neurogenet. 2018 May 2:1-12. PMID: 29718742.

From the abstract: "Despite availability of a diverse range of anti-epileptic drugs (AEDs), only about two-thirds of epilepsy patients respond well to drug treatment. Thus, novel targets are required to catalyse the design of next-generation AEDs. Manipulation of neuron firing-rate homoeostasis, through enhancing Pumilio (Pum) activity, has been shown to be potently anticonvulsant in Drosophila. In this study, we performed a genome-wide RNAi screen in S2R + cells, using a luciferase-based dPum activity reporter and identified 1166 genes involved in dPum regulation. Of these genes, we focused on 699 genes that, on knock-down, potentiate dPum activity/expression. Of this subgroup, 101 genes are activity-dependent based on comparison with genes previously identified as activity-dependent by RNA-sequencing. ... To test for anticonvulsant activity, we utilised an RNA-interference approach in vivo. RNAi-mediated knockdown showed that 57/101 genes (61%) are sufficient to significantly reduce seizure duration in the characterized seizure mutant, para[bss]. ... this study validates Pum as a favourable target for AED design and, moreover, identifies a number of lead compounds capable of increasing the expression of this homeostatic regulator."

Tuesday, March 6, 2018

FlyBook review on RNAi screening in Drosophila cells and in vivo

Heigwer F, Port F, Boutros M. RNA Interference (RNAi) Screening in Drosophila. Genetics. 2018 Mar;208(3):853-874. PMID: 29487145.

From the abstract: "... RNA interference (RNAi) ... has had an important impact on identifying and characterizing gene function. First discovered in Caenorhabditis elegans, RNAi can be used to silence the expression of genes through introduction of exogenous double-stranded RNA into cells. In Drosophila, RNAi has been applied in cultured cells or in vivo to perturb the function of single genes or to systematically probe gene function on a genome-wide scale. In this review, we will describe the use of RNAi to study gene function in Drosophila with a particular focus on high-throughput screening methods applied in cultured cells. ..."

Friday, February 23, 2018

Fly iPCs? Opinion piece in Genes suggests it should be tried

Kaur P, Jin HJ, Lusk JB, Tolwinski NS. Modeling the Role of Wnt Signaling in Human and Drosophila Stem Cells. Genes (Basel). 2018 Feb 16;9(2). pii: E101. PMID: 29462894.

From the abstract: "The discovery of induced pluripotent stem (iPS) cells ... dramatically transformed the study of stem cells ... Although advances have pushed the field forward, human application remains some years away, in part due to the need for an in-depth mechanistic understanding. The role of Wnts in stem cells predates the discovery of iPS cells with Wnts established as major pluripotency promoting factors. Most work to date has been done using mouse and tissue culture models and few attempts have been made in other model organisms, but the recent combination of clustered regularly interspaced short palindromic repeats (CRISPR) gene editing with iPS cell technology provides a perfect avenue for exploring iPS cells in model organisms. ... In this opinion article, we draw parallels between Wnt signaling in human and Drosophila stem cell systems, propose ways to obtain Drosophila iPS cells, and suggest ways to exploit the versatility of the Drosophila system for future stem cell studies."

Of related interest: RasV12 method for isolation of new cell lines, developed by A. Simcox (OSU).