High-throughput data analysis

2018 Apr 13

DRSC & TRiP Workshop at ADRC

1:45pm to 3:45pm

Location: 

Philadelphia, PA, USA
The DRSC & TRiP will be hosting a workshop at the Annual Drosophila Research Conference in Philadelphia, PA. The workshop is scheduled for Friday, April 13th from 1:45 to 3:45 PM. Come hear from DRSC & TRiP leaders Norbert Perrimon, Jonathan Zirin (organizer), Claire Yanhui Hu, and Stephanie Mohr. At the workshop, you will learn about new opportunities for community nomination and experiments using CRISPR knockout and activation, as well as learn what's new and popular among our online software and database tools. There will be something for everyone -- we will provide information... Read more about DRSC & TRiP Workshop at ADRC
Figure 2 from Housden et al 2017 PNAS

Variable Dose Analysis: a new DRSC-supported cell screen approach that leverages existing reagents to perform robust screens

December 1, 2017

We are excited to report the publication of a paper from Benjamin Housden and colleagues describing development and use of the Variable Dose Analysis (VDA) approach. Ben developed a way to use existing TRiP shRNA plasmids originally developed for fly stock production in a new, effective approach to high-throughput cell screening.

The VDA approach is particularly useful for combinatorial approaches that are acutely sensitive to assay robustness. The screen Ben and colleagues report focused on synthetic effects in Drosophila tumor model cells.  The...

Read more about Variable Dose Analysis: a new DRSC-supported cell screen approach that leverages existing reagents to perform robust screens
Screenshot of a 2015 Science paper from Payre and colleagues

Francois Payre's plenary talk at ADRC 2017 features results from DRSC cell-based screen

March 30, 2017

Those of us lucky enough to be at the Annual Drosophila Research Conference this morning saw a great talk by Francois Payre about regulation of Shavenbaby by small ORFs. A genome-wide cell-based screen done at the DRSC by Emilie Benrabah identified the mechanism of regulation. As this exemplifies, cell screens can help identify key pathways and factors that can then be followed up with in vivo studies.

J Zanet, E Benrabah, T Li, A Pélissier-Monier, H Chanut-Delalande, B Ronsin, HJ Bellen, F Payre, and S Plaza. 2015. “Pri sORF peptides induce selective proteasome-mediated protein processing.” Science, 349, 6254, Pp. 1356-8.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. Upon interaction with Ubr3, Pri peptides promote the binding of Ubr3 to Svb. Ubr3 can then ubiquitinate the Svb N terminus, which is degraded by the proteasome. The C-terminal domains protect Svb from complete degradation and ensure appropriate processing. Our data show that Pri peptides control selectivity of Ubr3 binding, which suggests that the family of sORF peptides may contain an extended repertoire of protein regulators.

Arunachalam Vinayagam, Meghana M Kulkarni, Richelle Sopko, Xiaoyun Sun, Yanhui Hu, Ankita Nand, Christians Villalta, Ahmadali Moghimi, Xuemei Yang, Stephanie E Mohr, Pengyu Hong, John M Asara, and Norbert Perrimon. 9/13/2016. “An Integrative Analysis of the InR/PI3K/Akt Network Identifies the Dynamic Response to Insulin Signaling.” Cell Reports, 16, 11, Pp. 3062-3074.Abstract

Insulin regulates an essential conserved signaling pathway affecting growth, proliferation, and meta- bolism. To expand our understanding of the insulin pathway, we combine biochemical, genetic, and computational approaches to build a comprehensive Drosophila InR/PI3K/Akt network. First, we map the dynamic protein-protein interaction network sur- rounding the insulin core pathway using bait-prey interactions connecting 566 proteins. Combining RNAi screening and phospho-specific antibodies, we find that 47% of interacting proteins affect pathway activity, and, using quantitative phospho- proteomics, we demonstrate that $10% of interact- ing proteins are regulated by insulin stimulation at the level of phosphorylation. Next, we integrate these orthogonal datasets to characterize the structure and dynamics of the insulin network at the level of protein complexes and validate our method by iden- tifying regulatory roles for the Protein Phosphatase 2A (PP2A) and Reptin-Pontin chromatin-remodeling complexes as negative and positive regulators of ribosome biogenesis, respectively. Altogether, our study represents a comprehensive resource for the study of the evolutionary conserved insulin network. 

Ian T Flockhart, Matthew Booker, Yanhui Hu, Benjamin McElvany, Quentin Gilly, Bernard Mathey-Prevot, Norbert Perrimon, and Stephanie E Mohr. 2012. “FlyRNAi.org--the database of the Drosophila RNAi screening center: 2012 update.” Nucleic Acids Res, 40, Database issue, Pp. D715-9.Abstract

FlyRNAi (http://www.flyrnai.org), the database and website of the Drosophila RNAi Screening Center (DRSC) at Harvard Medical School, serves a dual role, tracking both production of reagents for RNA interference (RNAi) screening in Drosophila cells and RNAi screen results. The database and website is used as a platform for community availability of protocols, tools, and other resources useful to researchers planning, conducting, analyzing or interpreting the results of Drosophila RNAi screens. Based on our own experience and user feedback, we have made several changes. Specifically, we have restructured the database to accommodate new types of reagents; added information about new RNAi libraries and other reagents; updated the user interface and website; and added new tools of use to the Drosophila community and others. Overall, the result is a more useful, flexible and comprehensive website and database.

Dawei Jiang, Linlin Zhao, and David E Clapham. 2009. “Genome-wide RNAi screen identifies Letm1 as a mitochondrial Ca2+/H+ antiporter.” Science, 326, 5949, Pp. 144-7.Abstract

Mitochondria are integral components of cellular calcium (Ca2+) signaling. Calcium stimulates mitochondrial adenosine 5'-triphosphate production, but can also initiate apoptosis. In turn, cytoplasmic Ca2+ concentrations are regulated by mitochondria. Although several transporter and ion-channel mechanisms have been measured in mitochondria, the molecules that govern Ca2+ movement across the inner mitochondrial membrane are unknown. We searched for genes that regulate mitochondrial Ca2+ and H+ concentrations using a genome-wide Drosophila RNA interference (RNAi) screen. The mammalian homolog of one Drosophila gene identified in the screen, Letm1, was found to specifically mediate coupled Ca2+/H+ exchange. RNAi knockdown, overexpression, and liposome reconstitution of the purified Letm1 protein demonstrate that Letm1 is a mitochondrial Ca2+/H+ antiporter.

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