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Drosophila chromatin

Laura Brueckner, Joris van Arensbergen, Waseem Akhtar, Ludo Pagie, Bas van Steensel
BACKGROUND: Chromatin proteins control gene activity in a concerted manner. We developed a high-throughput assay to study the effects of the local chromatin environment on the regulatory activity of a protein of interest. The assay combines a previously reported multiplexing strategy based on barcoded randomly integrated reporters with Gal4-mediated tethering. We applied the assay to Drosophila heterochromatin protein 1a (HP1a), which is mostly known as a repressive protein but has also been linked to transcriptional activation...
2016: Epigenetics & Chromatin
Daniil A Maksimov, Petr P Laktionov, Stepan N Belyakin
Analysis of gene expression regulation typically requires identification of genomic sites bound by regulatory proteins. For this purpose, chromatin immunoprecipitation (ChIP) and Dam identification (DamID) methods can be applied to cell lines, whole organisms, or enriched cell populations. In this work, we present modifications to the experimental DamID protocol, as well as a custom data processing algorithm, that allow to confidently identify genomic sites enriched with the proteins of interest. This algorithm is implemented in Perl and is also available as executable files, thereby making DamID analysis relatively straightforward...
October 21, 2016: Chromosome Research
Yad Ghavi-Helm, Bingqing Zhao, Eileen E M Furlong
Chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq) is an invaluable technique to assess transcription factor binding and histone modifications in a genome-wide manner, an essential step towards understanding the mechanisms that govern embryonic development. Here, we provide a detailed protocol for all steps involved in generating a ChIP-seq library, starting from embryo collection, fixation, chromatin preparation, immunoprecipitation, and finally library preparation. The protocol is optimized for Drosophila embryos, but can be easily adapted for any model organism...
2016: Methods in Molecular Biology
Mitzi I Kuroda, Andres Hilfiker, John C Lucchesi
The sex chromosomes have special significance in the history of genetics. The chromosomal basis of inheritance was firmly established when Calvin Bridges demonstrated that exceptions to Mendel's laws of segregation were accompanied at the cytological level by exceptional sex chromosome segregation. The morphological differences between X and Y exploited in Bridges' experiments arose as a consequence of the evolution of the sex chromosomes. Originally a homologous chromosome pair, the degeneration of the Y chromosome has been accompanied by a requirement for increased expression of the single X chromosome in males...
October 2016: Genetics
Lucia Mendoza-Viveros, Pascale Bouchard-Cannon, Sara Hegazi, Arthur H Cheng, Stephen Pastore, Hai-Ying Mary Cheng
Circadian timekeeping is a ubiquitous mechanism that enables organisms to maintain temporal coordination between internal biological processes and time of the local environment. The molecular basis of circadian rhythms lies in a set of transcription-translation feedback loops (TTFLs) that drives the rhythmic transcription of core clock genes, whose level and phase of expression serve as the marker of circadian time. However, it has become increasingly evident that additional regulatory mechanisms impinge upon the TTFLs to govern the properties and behavior of the circadian clock...
September 29, 2016: Cellular and Molecular Life Sciences: CMLS
Sung-Tae Hong, Kwang-Wook Choi
Genome stability is essential for all organisms. Translationally controlled tumour protein (TCTP) is a conserved protein associated with cancers. TCTP is involved in multiple intracellular functions, but its role in transcription and genome stability is poorly understood. Here, we demonstrate new functions of Drosophila TCTP (Tctp) in transcription and the stability of repeated sequences (rDNA and pericentromeric heterochromatin). Tctp binds Brahma (Brm) chromatin remodeler to negatively modulate its activity...
2016: Nature Communications
Na Xu, Xingwu Lu, Harsh Kavi, Alexander V Emelyanov, Travis J Bernardo, Elena Vershilova, Arthur I Skoultchi, Dmitry V Fyodorov
Metazoan linker histones are essential for development and play crucial roles in organization of chromatin, modification of epigenetic states and regulation of genetic activity. Vertebrates express multiple linker histone H1 isoforms, which may function redundantly. In contrast, H1 isoforms are not present in Dipterans, including D. melanogaster, except for an embryo-specific, distantly related dBigH1. Here we show that Drosophila BEN domain protein Elba2, which is expressed in early embryos and was hypothesized to have insulator-specific functions, can compensate for the loss of H1 in vivo...
September 30, 2016: Scientific Reports
Nina Koenecke, Jeff Johnston, Bjoern Gaertner, Malini Natarajan, Julia Zeitlinger
BACKGROUND: Drosophila dorso-ventral (DV) patterning is one of the best-understood regulatory networks to date, and illustrates the fundamental role of enhancers in controlling patterning, cell fate specification, and morphogenesis during development. Histone acetylation such as H3K27ac is an excellent marker for active enhancers, but it is challenging to obtain precise locations for enhancers as the highest levels of this modification flank the enhancer regions. How to best identify tissue-specific enhancers in a developmental system de novo with a minimal set of data is still unclear...
September 27, 2016: Genome Biology
Amanda Swain, Ziva Misulovin, Michelle Pherson, Maria Gause, Kathie Mihindukulasuriya, Ryan A Rickels, Ali Shilatifard, Dale Dorsett
The cohesin protein complex mediates sister chromatid cohesion and participates in transcriptional control of genes that regulate growth and development. Substantial reduction of cohesin activity alters transcription of many genes without disrupting chromosome segregation. Drosophila Nipped-B protein loads cohesin onto chromosomes, and together Nipped-B and cohesin occupy essentially all active transcriptional enhancers and a large fraction of active genes. It is unknown why some active genes bind high levels of cohesin and some do not...
September 2016: PLoS Genetics
Frédéric Bantignies
Initially discovered as repressors of homeotic gene expression in Drosophila, Polycomb group (PcG) proteins have now been shown to be involved in a plethora of biological processes. Indeed, by repressing a large number of target genes, including specific lineage genes, these chromatin factors play major roles in a multitude of cellular functions, such as pluripotency, differentiation, reprogramming, tissue regeneration, and nuclear organization. In this book chapter are presented in vivo approaches and technologies, which have been used in both mammalian and Drosophila systems to study the cellular functions of Polycomb group proteins...
2016: Methods in Molecular Biology
Benjamin Leblanc, Itys Comet, Frédéric Bantignies, Giacomo Cavalli
4C methods are useful to investigate dependencies between regulatory mechanisms and chromatin structures by revealing the frequency of chromatin contacts between a locus of interest and remote sequences on the chromosome. In this chapter we describe a protocol for the data analysis of microarray-based 4C experiments, presenting updated versions of the methods we used in a previous study of the large-scale chromatin interaction profile of a Polycomb response element in Drosophila. The protocol covers data preparation, normalization, microarray probe selection, and the multi-resolution detection of regions with enriched chromatin contacts...
2016: Methods in Molecular Biology
Hua-Bing Li
Linear chromatin fiber is packed inside the nuclei as a complex three-dimensional structure, and the organization of the chromatin has important roles in the appropriate spatial and temporal regulation of gene expression. To understand how chromatin organizes inside nuclei, and how regulatory proteins physically interact with genes, chromosome conformation capture (3C) technique provides a powerful and sensitive tool to detect both short- and long-range DNA-DNA interaction. Here I describe the 3C technique to detect the DNA-DNA interactions mediated by insulator proteins that are closely related to PcG in Drosophila, which is also broadly applicable to other systems...
2016: Methods in Molecular Biology
Federica Marasca, Fabrizia Marullo, Chiara Lanzuolo
Epigenetic mechanisms modulate and maintain the transcriptional state of the genome acting at various levels on chromatin. Emerging findings suggest that the position in the nuclear space and the cross talk between components of the nuclear architecture play a role in the regulation of epigenetic signatures. We recently described a cross talk between the Polycomb group of proteins (PcG) epigenetic repressors and the nuclear lamina. This interplay is important for the maintenance of transcriptional repression at muscle-specific genes and for the correct timing of muscle differentiation...
2016: Methods in Molecular Biology
Eva Löser, Daniel Latreille, Nicola Iovino
This protocol provides specific details on how to perform Chromatin immunoprecipitation (ChIP) from Drosophila embryos. ChIP allows the matching of proteins or histone modifications to specific genomic regions. Formaldehyde-cross-linked chromatin is isolated and antibodies against the target of interest are used to determine whether the target is associated with a specific DNA sequence. This can be performed in spatial and temporal manner and it can provide information about the genome-wide localization of a given protein or histone modification if coupled with deep sequencing technology (ChIP-Seq)...
2016: Methods in Molecular Biology
Pim van der Harst, Jessica van Setten, Niek Verweij, Georg Vogler, Lude Franke, Matthew T Maurano, Xinchen Wang, Irene Mateo Leach, Mark Eijgelsheim, Nona Sotoodehnia, Caroline Hayward, Rossella Sorice, Osorio Meirelles, Leo-Pekka Lyytikäinen, Ozren Polašek, Toshiko Tanaka, Dan E Arking, Sheila Ulivi, Stella Trompet, Martina Müller-Nurasyid, Albert V Smith, Marcus Dörr, Kathleen F Kerr, Jared W Magnani, Fabiola Del Greco M, Weihua Zhang, Ilja M Nolte, Claudia T Silva, Sandosh Padmanabhan, Vinicius Tragante, Tõnu Esko, Gonçalo R Abecasis, Michiel E Adriaens, Karl Andersen, Phil Barnett, Joshua C Bis, Rolf Bodmer, Brendan M Buckley, Harry Campbell, Megan V Cannon, Aravinda Chakravarti, Lin Y Chen, Alessandro Delitala, Richard B Devereux, Pieter A Doevendans, Anna F Dominiczak, Luigi Ferrucci, Ian Ford, Christian Gieger, Tamara B Harris, Eric Haugen, Matthias Heinig, Dena G Hernandez, Hans L Hillege, Joel N Hirschhorn, Albert Hofman, Norbert Hubner, Shih-Jen Hwang, Annamaria Iorio, Mika Kähönen, Manolis Kellis, Ivana Kolcic, Ishminder K Kooner, Jaspal S Kooner, Jan A Kors, Edward G Lakatta, Kasper Lage, Lenore J Launer, Daniel Levy, Alicia Lundby, Peter W Macfarlane, Dalit May, Thomas Meitinger, Andres Metspalu, Stefania Nappo, Silvia Naitza, Shane Neph, Alex S Nord, Teresa Nutile, Peter M Okin, Jesper V Olsen, Ben A Oostra, Josef M Penninger, Len A Pennacchio, Tune H Pers, Siegfried Perz, Annette Peters, Yigal M Pinto, Arne Pfeufer, Maria Grazia Pilia, Peter P Pramstaller, Bram P Prins, Olli T Raitakari, Soumya Raychaudhuri, Ken M Rice, Elizabeth J Rossin, Jerome I Rotter, Sebastian Schafer, David Schlessinger, Carsten O Schmidt, Jobanpreet Sehmi, Herman H W Silljé, Gianfranco Sinagra, Moritz F Sinner, Kamil Slowikowski, Elsayed Z Soliman, Timothy D Spector, Wilko Spiering, John A Stamatoyannopoulos, Ronald P Stolk, Konstantin Strauch, Sian-Tsung Tan, Kirill V Tarasov, Bosco Trinh, Andre G Uitterlinden, Malou van den Boogaard, Cornelia M van Duijn, Wiek H van Gilst, Jorma S Viikari, Peter M Visscher, Veronique Vitart, Uwe Völker, Melanie Waldenberger, Christian X Weichenberger, Harm-Jan Westra, Cisca Wijmenga, Bruce H Wolffenbuttel, Jian Yang, Connie R Bezzina, Patricia B Munroe, Harold Snieder, Alan F Wright, Igor Rudan, Laurie A Boyer, Folkert W Asselbergs, Dirk J van Veldhuisen, Bruno H Stricker, Bruce M Psaty, Marina Ciullo, Serena Sanna, Terho Lehtimäki, James F Wilson, Stefania Bandinelli, Alvaro Alonso, Paolo Gasparini, J Wouter Jukema, Stefan Kääb, Vilmundur Gudnason, Stephan B Felix, Susan R Heckbert, Rudolf A de Boer, Christopher Newton-Cheh, Andrew A Hicks, John C Chambers, Yalda Jamshidi, Axel Visel, Vincent M Christoffels, Aaron Isaacs, Nilesh J Samani, Paul I W de Bakker
BACKGROUND: Myocardial mass is a key determinant of cardiac muscle function and hypertrophy. Myocardial depolarization leading to cardiac muscle contraction is reflected by the amplitude and duration of the QRS complex on the electrocardiogram (ECG). Abnormal QRS amplitude or duration reflect changes in myocardial mass and conduction, and are associated with increased risk of heart failure and death. OBJECTIVES: This meta-analysis sought to gain insights into the genetic determinants of myocardial mass...
September 27, 2016: Journal of the American College of Cardiology
Toshiyuki Harumoto, Hisashi Anbutsu, Bruno Lemaitre, Takema Fukatsu
Some symbiotic bacteria are capable of interfering with host reproduction in selfish ways. How such bacteria can manipulate host's sex-related mechanisms is of fundamental interest encompassing cell, developmental and evolutionary biology. Here, we uncover the molecular and cellular mechanisms underlying Spiroplasma-induced embryonic male lethality in Drosophila melanogaster. Transcriptomic analysis reveals that many genes related to DNA damage and apoptosis are up-regulated specifically in infected male embryos...
September 21, 2016: Nature Communications
Youqiong Ye, Liang Gu, Xiaolong Chen, Jiejun Shi, Xiaobai Zhang, Cizhong Jiang
Chromatin remodeling plays a critical role in gene regulation and impacts many biological processes. However, little is known about the relationship between chromatin remodeling dynamics and in vivo cell lineage commitment. Here, we reveal the patterns of histone modification change and nucleosome positioning dynamics and their epigenetic regulatory roles during the in vivo glial differentiation in early Drosophila embryos. The genome-wide average H3K9ac signals in promoter regions are decreased in the glial cells compared to the neural progenitor cells...
2016: Scientific Reports
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, Norbert Perrimon
Insulin regulates an essential conserved signaling pathway affecting growth, proliferation, and metabolism. 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 surrounding 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 phosphoproteomics, we demonstrate that ∼10% of interacting proteins are regulated by insulin stimulation at the level of phosphorylation...
September 13, 2016: Cell Reports
S V Satya Prakash Avva, Craig M Hart
Data implicate the Drosophila 32 kDa Boundary Element-Associated Factors BEAF-32A and BEAF-32B in both chromatin domain insulator element function and promoter function. They might also function as an epigenetic memory by remaining bound to mitotic chromosomes. Both proteins are made from the same gene. They differ in their N-terminal 80 amino acids, which contain single DNA-binding BED fingers. The remaining 200 amino acids are identical in the two proteins. The structure and function of the middle region of 120 amino acids is unknown, while the C-terminal region of 80 amino acids has a putative leucine zipper and a BESS domain and mediates BEAF-BEAF interactions...
2016: PloS One
Zhibo Ma, Mo Li, Sharmila Roy, Kevin J Liu, Matthew L Romine, Derrick C Lane, Sapna K Patel, Haini N Cai
The three-dimensional (3D) organization of the eukaryotic genome is critical for its proper function. Evidence suggests that extensive chromatin loops form the building blocks of the genomic architecture, separating genes and gene clusters into distinct functional domains. These loops are anchored in part by a special type of DNA elements called chromatin boundary elements (CBEs). CBEs were originally found to insulate neighboring genes by blocking influences of transcriptional enhancers or the spread of silent chromatin...
August 26, 2016: World Journal of Biological Chemistry
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