Genome-wide RNAi Screen Identifies Networks Involved in Intestinal Stem Cell Regulation in Drosophila

Zeng, Xiankun; Han, Li‐Li; Singh, Shree Ram; Liu, Hanhan; Neumüller, Ralph A.; Dong, Yan; Hu, Yanhui; Liu, Ying; Liu, Wei; Lin, Xinhua; Hou, Steven X.

doi:10.1016/j.celrep.2015.01.051

Cited by 93 publications

(117 citation statements)

References 71 publications

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“…In particular, results from various large-scale screens have been included. Among these are germline and maternal effect screens using the MTD-Gal4 driver (Staller et al 2013;Sopko et al 2014;Yan et al 2014), a muscle screen using the muscle Dmef2-Gal4 line (Perrimon and Randkelv, unpublished results), a screen in gut stem cells using the esg-Gal4 (Zeng et al 2015), and a screen of maternally expressed genes involved in embryonic patterning (Liu and Lasko 2015). As of May 2015, and specific for the TRiP stocks, the RSVP contains 8334 data entries for 5202 TRiP stocks representing 3735 fly genes.…”

Section: Validation Of the Trip Linesmentioning

confidence: 99%

The Transgenic RNAi Project at Harvard Medical School: Resources and Validation

et al. 2015

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To facilitate large-scale functional studies in Drosophila, the Drosophila Transgenic RNAi Project (TRiP) at Harvard Medical School (HMS) was established along with several goals: developing efficient vectors for RNAi that work in all tissues, generating a genome-scale collection of RNAi stocks with input from the community, distributing the lines as they are generated through existing stock centers, validating as many lines as possible using RT-qPCR and phenotypic analyses, and developing tools and web resources for identifying RNAi lines and retrieving existing information on their quality. With these goals in mind, here we describe in detail the various tools we developed and the status of the collection, which is currently composed of 11,491 lines and covering 71% of Drosophila genes. Data on the characterization of the lines either by RT-qPCR or phenotype is available on a dedicated website, the RNAi Stock Validation and Phenotypes Project (RSVP, http://www.flyrnai.org/RSVP.html), and stocks are available from three stock centers, the Bloomington Drosophila Stock Center (United States), National Institute of Genetics (Japan), and TsingHua Fly Center (China). KEYWORDS RNAi; Drosophila; screens; phenotypes; functional genomics A striking finding from the genomic revolution and wholegenome sequencing is the amount of information missing on gene function. Although Drosophila is arguably the bestunderstood multicellular organism and a proven model system for human diseases, mutations mapped to specific genes with readily detectable phenotypes have been isolated for 15% of the .13919 annotated fly coding genes (http:// flybase.org/; FlyBase R6.06). The lack of information on the majority of genes (the "phenotype gap") suggests that researchers have been unable to either assay their roles experimentally and/or resolve issues of functional redundancy. In addition, some phenotypes may be only detected on specific diets and environments. Further, our understanding of the function of many genes for which we have some information is limited by pleiotropy, whereby an earlier function of the gene prevents analysis of later functions.The availability of in vivo RNAi has revolutionized the ability of Drosophila researchers to disrupt the activity of single genes with spatial and temporal resolution (Dietzl et al. 2007; see review by Perrimon et al. 2010), and thus address the phenotype gap. Motivated by the power of the approach and the needs of the community, three large-scale efforts, the Vienna Drosophila RNAi Center (VDRC, http:// stockcenter.vdrc.at/control/main), the National Institute of Genetics (NIG, http://www.shigen.nig.ac.jp/fly/nigfly/index.jsp), and the Drosophila Transgenic RNAi Project (TRiP) at Harvard Medical School (HMS) (http://www.flyrnai.org/TRiP-HOME. html) have over the years generated large numbers of RNAi lines that aim to cover all Drosophila genes. These resources are proving invaluable to address a myriad of questions in various biological and biomedical fields including but not limite...

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Section: Validation Of the Trip Linesmentioning

confidence: 99%

The Transgenic RNAi Project at Harvard Medical School: Resources and Validation

et al. 2015

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“…Several RNAi screens that have used the same collection of RNAi lines suggest that the off-target rates of this collection are low (Ni et al, 2011;Yan et al, 2014;Zeng et al, 2015). Of the 211 genes identified here, 31 turned out to have two independent shRNAi lines.…”

Section: Gsc Self-renewal Screen In Fly Testesmentioning

confidence: 99%

“…Thus far, several large-scale RNAi screens have been carried out in various types of stem cells to identify the regulatory networks of self-renewal and differentiation. We therefore compared our screened data with data sets obtained from other stem cell-related screens to identify common or unique factors that regulate self-renewal and differentiation of different stem cells (Neumuller et al, 2011;Yan et al, 2014;Zeng et al, 2015).…”

Section: Non-cell Autonomous Effects Of Germ Cell Differentiationmentioning

confidence: 99%

Protein synthesis and degradation are essential to regulate germline stem cell homeostasis in Drosophila testes

Xiang

Chen

et al. 2016

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The homeostasis of self-renewal and differentiation in stem cells is controlled by intrinsic signals and their niche. We conducted a large-scale RNA interference (RNAi) screen in Drosophila testes and identified 221 genes required for germline stem cell (GSC) maintenance or differentiation. Knockdown of these genes in transit-amplifying spermatogonia and cyst cells further revealed various phenotypes. Complex analysis uncovered that many of the identified genes are involved in key steps of protein synthesis and degradation. A group of genes that are required for mRNA splicing and protein translation contributes to both GSC selfrenewal and early germ cell differentiation. Loss of genes in the protein degradation pathway in cyst cells leads to testis tumors consisting of overproliferated germ cells. Importantly, in the Cullin 4-RING E3 ubiquitin ligase (CRL4) complex, we identified multiple proteins that are crucial to GSC self-renewal: pic/DDB1, a CRL4 linker protein, is not only required for GSC self-renewal in flies but also for maintenance of spermatogonial stem cells (SSCs) in mice.

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“…3A-C). Slit/Robo signaling was recently identified as a negative regulator of EE fate commitment (Biteau and Jasper, 2014;Zeng et al, 2015). Atg16 expression was lower in stem cells positive for Robo2 and Delta ( Fig.…”

Section: Isc Niche Signaling Is Altered In Atg16 Wd40 Mutantsmentioning

confidence: 99%

“…Fully differentiated EE cells secrete Slit, which acts on its receptor Robo2 (also known as Leak in Drosophila) that is expressed by both ISCs and differentiating precursor cells (Biteau and Jasper, 2014;Zeng et al, 2015). Activation of Robo signaling represses the homeodomain transcription factor Prospero, leading to the differentiation of EBs into EC instead of EE cells.…”

Section: Introductionmentioning

confidence: 99%

Drosophila Atg16 promotes enteroendocrine cell differentiation via regulation of intestinal Slit/Robo signaling

et al. 2017

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Genetic variations of Atg16l1, Slit2 and Rab19 predispose to the development of inflammatory bowel disease (IBD), but the relationship between these mutations is unclear. Here we show that in Drosophila guts lacking the WD40 domain of Atg16, pre-enteroendocrine (pre-EE) cells accumulate that fail to differentiate into properly functioning secretory EE cells. Mechanistically, loss of Atg16 or its binding partner Rab19 impairs Slit production, which normally inhibits EE cell generation by activating Robo signaling in stem cells. Importantly, loss of Atg16 or decreased Slit/Robo signaling triggers an intestinal inflammatory response. Surprisingly, analysis of Rab19 and domainspecific Atg16 mutants indicates that their stem cell niche regulatory function is independent of autophagy. Our study reveals how mutations in these different genes may contribute to IBD.

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Genome-wide RNAi Screen Identifies Networks Involved in Intestinal Stem Cell Regulation in Drosophila

Cited by 93 publications

References 71 publications

The Transgenic RNAi Project at Harvard Medical School: Resources and Validation

The Transgenic RNAi Project at Harvard Medical School: Resources and Validation

Protein synthesis and degradation are essential to regulate germline stem cell homeostasis in Drosophila testes

Drosophila Atg16 promotes enteroendocrine cell differentiation via regulation of intestinal Slit/Robo signaling

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