A genome-wide resource for the analysis of protein localisation in Drosophila

The Drosophila genome contains >13,000 protein coding genes, the majority of which remain poorly investigated. Important reasons include the lack of antibodies or reporter constructs to visualise these proteins. Here we present a genome-wide fosmid library of ≈10,000 GFP-tagged clones, comprising tagged genes and most of their regulatory information. For 880 tagged proteins we have created transgenic lines and for a total of 207 lines we have assessed protein expression and localisation in ovaries, embryos, pupae or adults by stainings and live imaging approaches. Importantly, we can visualise many proteins at endogenous expression levels and find a large fraction of them localising to subcellular compartments. Using complementation tests we demonstrate that two-thirds of the tagged proteins are fully functional. Moreover, our clones also enable interaction proteomics from developing pupae and adult flies. Taken together, this resource will enable systematic analysis of protein expression and localisation in various cellular and developmental contexts.

[1]  Tobias Pietzsch,et al.  An automated workflow for parallel processing of large multiview SPIM recordings , 2015, Bioinform..

[2]  Marco Y. Hein,et al.  A Human Interactome in Three Quantitative Dimensions Organized by Stoichiometries and Abundances , 2015, Cell.

[3]  Jaison J. Omoto,et al.  Lineage-associated tracts defining the anatomy of the Drosophila first instar larval brain. , 2015, Developmental biology.

[4]  A. Horovitz,et al.  Thermodynamic Protein Destabilization by GFP Tagging: A Case of Interdomain Allostery. , 2015, Biophysical journal.

[5]  Andrew G. York,et al.  Drosophila germ granules are structured and contain homotypic mRNA clusters , 2015, Nature Communications.

[6]  Tobias Pietzsch,et al.  BigDataViewer: visualization and processing for large image data sets , 2015, Nature Methods.

[7]  P. Tomançak,et al.  Endogenously Tagged Rab Proteins: A Resource to Study Membrane Trafficking in Drosophila , 2015, Developmental cell.

[8]  Benjamin W Booth,et al.  A library of MiMICs allows tagging of genes and reversible, spatial and temporal knockdown of proteins in Drosophila , 2015, eLife.

[9]  Michael F. Wangler,et al.  Fruit Flies in Biomedical Research , 2015, Genetics.

[10]  E. van Nimwegen,et al.  A large-scale, in vivo transcription factor screen defines bivalent chromatin as a key property of regulatory factors mediating Drosophila wing development , 2015, Genome research.

[11]  B. Habermann,et al.  The RNA-binding protein Arrest (Bruno) regulates alternative splicing to enable myofibril maturation in Drosophila flight muscle , 2014, EMBO reports.

[12]  Alex T. Kalinka,et al.  Systematic imaging reveals features and changing localization of mRNAs in Drosophila development , 2014, bioRxiv.

[13]  Marco Y. Hein,et al.  Accurate Protein Complex Retrieval by Affinity Enrichment Mass Spectrometry (AE-MS) Rather than Affinity Purification Mass Spectrometry (AP-MS)* , 2014, Molecular & Cellular Proteomics.

[14]  Dominique A. Glauser,et al.  A Conserved Role for p48 Homologs in Protecting Dopaminergic Neurons from Oxidative Stress , 2014, PLoS genetics.

[15]  Matthias Landgraf,et al.  Analysis of the expression patterns, subcellular localisations and interaction partners of Drosophila proteins using a pigP protein trap library , 2014, Development.

[16]  R. Cripps,et al.  Arrest is a regulator of fiber-specific alternative splicing in the indirect flight muscles of Drosophila , 2014, The Journal of cell biology.

[17]  Marco Y. Hein A human interactome , 2014 .

[18]  F. Schnorrer,et al.  A Versatile Two-Step CRISPR- and RMCE-Based Strategy for Efficient Genome Engineering in Drosophila , 2014, G3: Genes, Genomes, Genetics.

[19]  Simon L. Bullock,et al.  Optimized CRISPR/Cas tools for efficient germline and somatic genome engineering in Drosophila , 2014, Proceedings of the National Academy of Sciences.

[20]  Marco Y. Hein,et al.  Accurate Proteome-wide Label-free Quantification by Delayed Normalization and Maximal Peptide Ratio Extraction, Termed MaxLFQ * , 2014, Molecular & Cellular Proteomics.

[21]  F. Schnorrer,et al.  A guide to study Drosophila muscle biology. , 2014, Methods.

[22]  Danny E. Miller,et al.  Corolla Is a Novel Protein That Contributes to the Architecture of the Synaptonemal Complex of Drosophila , 2014, Genetics.

[23]  B. Dickson,et al.  Genome-scale functional characterization of Drosophila developmental enhancers in vivo , 2014, Nature.

[24]  A. Ephrussi,et al.  A stem–loop structure directs oskar mRNA to microtubule minus ends , 2014, RNA.

[25]  Frank Schnorrer,et al.  Tension and Force-Resistant Attachment Are Essential for Myofibrillogenesis in Drosophila Flight Muscle , 2014, Current Biology.

[26]  S. Vernes Genome wide identification of Fruitless targets suggests a role in upregulating genes important for neural circuit formation , 2014, Scientific Reports.

[27]  C. Rubinstein,et al.  Highly Specific and Efficient CRISPR/Cas9-Catalyzed Homology-Directed Repair in Drosophila , 2014, Genetics.

[28]  F. Schnorrer,et al.  Transcriptional regulation and alternative splicing cooperate in muscle fiber-type specification in flies and mammals , 2013, Experimental cell research.

[29]  Stephan Preibisch,et al.  Efficient Bayesian-based multiview deconvolution , 2013, Nature Methods.

[30]  Pavel Tomancak,et al.  Open-source solutions for SPIMage processing. , 2014, Methods in cell biology.

[31]  Manolis Kellis,et al.  Spatial expression of transcription factors in Drosophila embryonic organ development , 2013, Genome Biology.

[32]  Jaison J. Omoto,et al.  Postembryonic lineages of the Drosophila brain: I. Development of the lineage-associated fiber tracts. , 2013, Developmental biology.

[33]  C. Alexandre,et al.  Accelerated homologous recombination and subsequent genome modification in Drosophila , 2013, Development.

[34]  M. Noll,et al.  Differential and redundant functions of gooseberry and gooseberry neuro in the central nervous system and segmentation of the Drosophila embryo. , 2013, Developmental biology.

[35]  D. Johnston Using mutants, knockdowns, and transgenesis to investigate gene function in Drosophila. , 2013 .

[36]  J. Taipale,et al.  A versatile platform for creating a comprehensive UAS-ORFeome library in Drosophila , 2013, Development.

[37]  Emma Lundberg,et al.  Immunofluorescence and fluorescent-protein tagging show high correlation for protein localization in mammalian cells , 2013, Nature Methods.

[38]  Michael J. Hoffmann,et al.  big bang gene modulates gut immune tolerance in Drosophila , 2013, Proceedings of the National Academy of Sciences.

[39]  D. St Johnston Using mutants, knockdowns, and transgenesis to investigate gene function in Drosophila , 2013, Wiley interdisciplinary reviews. Developmental biology.

[40]  M. Furuse,et al.  A novel protein complex, Mesh–Ssk, is required for septate junction formation in the Drosophila midgut , 2012, Journal of Cell Science.

[41]  Anthony A. Hyman,et al.  A Genome-Scale Resource for In Vivo Tag-Based Protein Function Exploration in C. elegans , 2012, Cell.

[42]  Gabor T. Marth,et al.  Haplotype-based variant detection from short-read sequencing , 2012, 1207.3907.

[43]  K. Leonard,et al.  The function of the M-line protein obscurin in controlling the symmetry of the sarcomere in the flight muscle of Drosophila , 2012, Journal of Cell Science.

[44]  Johannes E. Schindelin,et al.  Fiji: an open-source platform for biological-image analysis , 2012, Nature Methods.

[45]  Steven L Salzberg,et al.  Fast gapped-read alignment with Bowtie 2 , 2012, Nature Methods.

[46]  N. Perrimon,et al.  Stringent Analysis of Gene Function and Protein–Protein Interactions Using Fluorescently Tagged Genes , 2012, Genetics.

[47]  H. Bellen,et al.  Genome-wide manipulations of Drosophila melanogaster with transposons, Flp recombinase, and ΦC31 integrase. , 2012, Methods in molecular biology.

[48]  L. Rasmussen,et al.  Functional Genomics , 2012, Methods in Molecular Biology.

[49]  M. Affolter,et al.  Fluorescent fusion protein knockout mediated by anti-GFP nanobody , 2011, Nature Structural &Molecular Biology.

[50]  A. Ephrussi,et al.  Dimerization of oskar 3' UTRs promotes hitchhiking for RNA localization in the Drosophila oocyte. , 2011, RNA.

[51]  Hans-Ulrich Dodt,et al.  Spalt mediates an evolutionarily conserved switch to fibrillar muscle fate in insects , 2011, Nature.

[52]  B. Edgar,et al.  Intestinal stem cells in the adult Drosophila midgut. , 2011, Experimental cell research.

[53]  Julian Mintseris,et al.  A Protein Complex Network of Drosophila melanogaster , 2011, Cell.

[54]  Nele A. Haelterman,et al.  MiMIC: a highly versatile transposon insertion resource for engineering Drosophila melanogaster genes , 2011, Nature Methods.

[55]  Norbert Perrimon,et al.  A genome-scale shRNA resource for transgenic RNAi in Drosophila , 2011, Nature Methods.

[56]  A. Stewart,et al.  Recombineering, transfection, Western, IP and ChIP methods for protein tagging via gene targeting or BAC transgenesis. , 2011, Methods.

[57]  B. Graveley The developmental transcriptome of Drosophila melanogaster , 2010, Nature.

[58]  P. Tomançak,et al.  Recombination-mediated genetic engineering of large genomic DNA transgenes. , 2011, Methods in molecular biology.

[59]  E. Popodi,et al.  A Molecularly Defined Duplication Set for the X Chromosome of Drosophila melanogaster , 2010, Genetics.

[60]  S. Henikoff,et al.  A simple method for gene expression and chromatin profiling of individual cell types within a tissue. , 2010, Developmental cell.

[61]  A. Hyman,et al.  Quantitative proteomics combined with BAC TransgeneOmics reveals in vivo protein interactions , 2010, The Journal of cell biology.

[62]  Cornelia Schönbauer,et al.  Systematic genetic analysis of muscle morphogenesis and function in Drosophila , 2010, Nature.

[63]  A. Spradling,et al.  clueless, a conserved Drosophila gene required for mitochondrial subcellular localization, interacts genetically with parkin , 2009, Disease Models & Mechanisms.

[64]  E. Knust,et al.  The Crumbs complex: from epithelial-cell polarity to retinal degeneration , 2009, Journal of Cell Science.

[65]  Gonçalo R. Abecasis,et al.  The Sequence Alignment/Map format and SAMtools , 2009, Bioinform..

[66]  Pavel Tomancak,et al.  A toolkit for high-throughput, cross-species gene engineering in Drosophila , 2009, Nature Methods.

[67]  K. White,et al.  Versatile P(acman) BAC Libraries for Transgenesis Studies in Drosophila melanogaster , 2009, Nature Methods.

[68]  Stephan Saalfeld,et al.  Globally optimal stitching of tiled 3D microscopic image acquisitions , 2009, Bioinform..

[69]  M. Mann,et al.  MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification , 2008, Nature Biotechnology.

[70]  Jon Heales Open source solutions , 2008 .

[71]  M. Mannervik,et al.  Wollknäuel is required for embryo patterning and encodes the Drosophila ALG5 UDP-glucose:dolichyl-phosphate glucosyltransferase , 2008, Development.

[72]  P. Tomançak,et al.  Global Analysis of mRNA Localization Reveals a Prominent Role in Organizing Cellular Architecture and Function , 2007, Cell.

[73]  D. Stainier,et al.  Even fluorescence excitation by multidirectional selective plane illumination microscopy (mSPIM). , 2007, Optics letters.

[74]  G. Rubin,et al.  Global analysis of patterns of gene expression during Drosophila embryogenesis , 2007, Genome Biology.

[75]  B. Dickson,et al.  A genome-wide transgenic RNAi library for conditional gene inactivation in Drosophila , 2007, Nature.

[76]  M. Beckerle,et al.  The Drosophila muscle LIM protein, Mlp84B, cooperates with D-titin to maintain muscle structural integrity , 2007, Journal of Cell Science.

[77]  Frank Schnorrer,et al.  The transmembrane protein Kon-tiki couples to Dgrip to mediate myotube targeting in Drosophila. , 2007, Developmental cell.

[78]  Roger A Hoskins,et al.  The Carnegie Protein Trap Library: A Versatile Tool for Drosophila Developmental Studies , 2007, Genetics.

[79]  R. Hoskins,et al.  Exploring Strategies for Protein Trapping in Drosophila , 2007, Genetics.

[80]  Hugo J. Bellen,et al.  P[acman]: A BAC Transgenic Platform for Targeted Insertion of Large DNA Fragments in D. melanogaster , 2006, Science.

[81]  Miklós Erdélyi,et al.  A translation-independent role of oskar RNA in early Drosophila oogenesis , 2006, Development.

[82]  Volker Hartenstein,et al.  Neural Lineages of the Drosophila Brain: A Three-Dimensional Digital Atlas of the Pattern of Lineage Location and Projection at the Late Larval Stage , 2006, The Journal of Neuroscience.

[83]  Sean R. Collins,et al.  Global landscape of protein complexes in the yeast Saccharomyces cerevisiae , 2006, Nature.

[84]  N. Perrimon,et al.  Functional genomics reveals genes involved in protein secretion and Golgi organization , 2006, Nature.

[85]  A. Spradling,et al.  The adult Drosophila posterior midgut is maintained by pluripotent stem cells , 2006, Nature.

[86]  T. Terwilliger,et al.  Engineering and characterization of a superfolder green fluorescent protein , 2006, Nature Biotechnology.

[87]  Jan Huisken,et al.  Selective Plane Illumination Microscopy , 2006 .

[88]  A dominant negative form of Rac1 affects myogenesis of adult thoracic muscles in Drosophila. , 2005, Developmental biology.

[89]  M. Cutler,et al.  The Ras suppressor Rsu-1 binds to the LIM 5 domain of the adaptor protein PINCH1 and participates in adhesion-related functions. , 2005, Experimental cell research.

[90]  R. Chanet,et al.  Protein interaction mapping: a Drosophila case study. , 2005, Genome research.

[91]  R. Chanet,et al.  Protein interaction mapping: A Drosophila case study (vol 15, pg 376, 2005) , 2005 .

[92]  John R. Yates,et al.  The integrin effector PINCH regulates JNK activity and epithelial migration in concert with Ras suppressor 1 , 2004, The Journal of cell biology.

[93]  Markus Fischer,et al.  A brilliant monomeric red fluorescent protein to visualize cytoskeleton dynamics in Dictyostelium , 2004, FEBS letters.

[94]  F. Del Bene,et al.  Optical Sectioning Deep Inside Live Embryos by Selective Plane Illumination Microscopy , 2004, Science.

[95]  Joerg Betschinger,et al.  Sequential roles of Cdc42, Par-6, aPKC, and Lgl in the establishment of epithelial polarity during Drosophila embryogenesis. , 2004, Developmental cell.

[96]  J. Clayton,et al.  Expression and function of the Drosophila ACT88F actin isoform is not restricted to the indirect flight muscles , 2004, Journal of Muscle Research & Cell Motility.

[97]  James R. Knight,et al.  A Protein Interaction Map of Drosophila melanogaster , 2003, Science.

[98]  G. Technau,et al.  Molecular markers for identified neuroblasts in the developing brain of Drosophila , 2003, Development.

[99]  E. R. Gavis,et al.  Live Imaging of Endogenous RNA Reveals a Diffusion and Entrapment Mechanism for nanos mRNA Localization in Drosophila , 2003, Current Biology.

[100]  Gilles Laurent,et al.  painless, a Drosophila Gene Essential for Nociception , 2003, Cell.

[101]  R. Cantera,et al.  Synaptic activity modifies the levels of Dorsal and Cactus at the neuromuscular junction of Drosophila. , 2003, Journal of neurobiology.

[102]  V. Hartenstein,et al.  Early development of the Drosophila brain: IV. Larval neuropile compartments defined by glial septa , 2003, The Journal of comparative neurology.

[103]  M. Ashburner,et al.  Systematic determination of patterns of gene expression during Drosophila embryogenesis , 2002, Genome Biology.

[104]  I. Hariharan,et al.  An overexpression screen in Drosophila for genes that restrict growth or cell-cycle progression in the developing eye. , 2002, Genetics.

[105]  Gary D Bader,et al.  Systematic identification of protein complexes in Saccharomyces cerevisiae by mass spectrometry , 2002, Nature.

[106]  P. Bork,et al.  Functional organization of the yeast proteome by systematic analysis of protein complexes , 2002, Nature.

[107]  Nancy Paterson The Library , 1912, Leonardo.

[108]  X. Morin,et al.  A protein trap strategy to detect GFP-tagged proteins expressed from their endogenous loci in Drosophila , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[109]  N. Gay,et al.  Amphiphysin is necessary for organization of the excitation-contraction coupling machinery of muscles, but not for synaptic vesicle endocytosis in Drosophila. , 2001, Genes & development.

[110]  Yun Hua,et al.  A New Focal Adhesion Protein That Interacts with Integrin-Linked Kinase and Regulates Cell Adhesion and Spreading , 2001, The Journal of cell biology.

[111]  C. Thummel,et al.  Genetic analysis of the Drosophila 63F early puff. Characterization of mutations in E63-1 and maggie, a putative Tom22. , 2000, Genetics.

[112]  Stephen M. Mount,et al.  The genome sequence of Drosophila melanogaster. , 2000, Science.

[113]  P. Salvaterra,et al.  Dynamic visualization of nervous system in live Drosophila. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[114]  F. Kafatos,et al.  Regulation of the spalt/spalt-related gene complex and its function during sensory organ development in the Drosophila thorax. , 1999, Development.

[115]  P. Rørth Gal4 in the Drosophila female germline , 1998, Mechanisms of Development.

[116]  G. Rubin,et al.  Systematic gain-of-function genetics in Drosophila. , 1998, Development.

[117]  B. Wanner,et al.  Conditionally replicative and conjugative plasmids carrying lacZ alpha for cloning, mutagenesis, and allele replacement in bacteria. , 1996, Plasmid.

[118]  P. Macdonald,et al.  Translational regulation of oskar mRNA by Bruno, an ovarian RNA-binding protein, is essential , 1995, Cell.

[119]  R M Cripps,et al.  Recovery of dominant, autosomal flightless mutants of Drosophila melanogaster and identification of a new gene required for normal muscle structure and function. , 1994, Genetics.

[120]  T. Schüpbach,et al.  The drosophila dorsoventral patterning gene gurken produces a dorsally localized RNA and encodes a TGFα-like protein , 1993, Cell.

[121]  N. Patel,et al.  Analysis of the gooseberry locus in Drosophila embryos: gooseberry determines the cuticular pattern and activates gooseberry neuro. , 1993, Development.

[122]  J. Vigoreaux,et al.  Flightin, a novel myofibrillar protein of Drosophila stretch-activated muscles , 1993, The Journal of cell biology.

[123]  M. Bate,et al.  Development of the indirect flight muscles of Drosophila. , 1991, Development.

[124]  P. Macdonald,et al.  oskar mRNA is localized to the posterior pole of the Drosophila oocyte , 1991, Cell.

[125]  R. Lehmann,et al.  oskar organizes the germ plasm and directs localization of the posterior determinant nanos , 1991, Cell.

[126]  R. Renkawitz-Pohl,et al.  β3 tubulin expression characterizes the differentiating mesodermal germ layer during Drosophila embryogenesis , 1988 .

[127]  R. Renkawitz-Pohl,et al.  Beta 3 tubulin expression characterizes the differentiating mesodermal germ layer during Drosophila embryogenesis. , 1988, Development.

[128]  L. Fessler,et al.  Drosophila laminin: characterization and localization , 1987, The Journal of cell biology.

[129]  Stanley N Cohen,et al.  Partitioning of bacterial plasmids during cell division: a cis-acting locus that accomplishes stable plasmid inheritance , 1980, Cell.

[130]  従二 和彦 抗ガン薬-protein Complex , 1977 .

[131]  F. Warner Analysis of expression. , 1885 .