The Large Isoform of Drosophila melanogaster Heterochromatin Protein 2 Plays a Critical Role in Gene Silencing and Chromosome Structure

Heterochromatin is enriched for specific epigenetic factors including Heterochromatin Protein 1a (HP1a), and is essential for many organismal functions. To elucidate heterochromatin organization and regulation, we purified Drosophila melanogaster HP1a interactors, and performed a genome-wide RNAi screen to identify genes that impact HP1a levels or localization. The majority of the over four hundred putative HP1a interactors and regulators identified were previously unknown. We found that 13 of 16 tested candidates (83%) are required for gene silencing, providing a substantial increase in the number of identified components that impact heterochromatin properties. Surprisingly, image analysis revealed that although some HP1a interactors and regulators are broadly distributed within the heterochromatin domain, most localize to discrete subdomains that display dynamic localization patterns during the cell cycle. We conclude that heterochromatin composition and architecture is more spatially complex and dynamic than previously suggested, and propose that a network of subdomains regulates diverse heterochromatin functions.

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Dual roles for the Drosophila PI 4-kinase Four wheel drive in localizing Rab11 during cytokinesis

The Journal of Cell Biology ◽

10.1083/jcb.200908107 ◽

2009 ◽

Vol 187 (6) ◽

pp. 847-858 ◽

Cited By ~ 88

Author(s):

Gordon Polevoy ◽

Ho-Chun Wei ◽

Raymond Wong ◽

Zsofia Szentpetery ◽

Yeun Ju Kim ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Critical Role ◽

Successful Completion ◽

Dual Roles ◽

Recycling Endosome ◽

Golgi Membranes ◽

Wheel Drive ◽

Dividing Cells ◽

Four Wheel Drive

Successful completion of cytokinesis relies on addition of new membrane, and requires the recycling endosome regulator Rab11, which localizes to the midzone. Despite the critical role of Rab11 in this process, little is known about the formation and composition of Rab11-containing organelles. Here, we identify the phosphatidylinositol (PI) 4-kinase III β Four wheel drive (Fwd) as a key regulator of Rab11 during cytokinesis in Drosophila melanogaster spermatocytes. We show Fwd is required for synthesis of PI 4-phosphate (PI4P) on Golgi membranes and for formation of PI4P-containing secretory organelles that localize to the midzone. Fwd binds and colocalizes with Rab11 on Golgi membranes, and is required for localization of Rab11 in dividing cells. A kinase-dead version of Fwd also binds Rab11 and partially restores cytokinesis to fwd mutant flies. Moreover, activated Rab11 partially suppresses loss of fwd. Our data suggest Fwd plays catalytic and noncatalytic roles in regulating Rab11 during cytokinesis.

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The P-element-induced silencing effect of KP transposons is dose dependent in Drosophila melanogaster

Genome ◽

10.1139/g11-023 ◽

2011 ◽

Vol 54 (9) ◽

pp. 752-762 ◽

Cited By ~ 7

Author(s):

Alireza Sameny ◽

John Locke

Keyword(s):

Drosophila Melanogaster ◽

Critical Role ◽

Mutagenic Effect ◽

P Element ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

P Elements ◽

Single Well ◽

Dose Dependent

Transposable elements are found in the genomes of all eukaryotes and play a critical role in altering gene expression and genome organization. In Drosophila melanogaster, transposable P elements are responsible for the phenomenon of hybrid dysgenesis. KP elements, a deletion-derivative of the complete P element, can suppress this mutagenic effect. KP elements can also silence the expression of certain other P-element-mediated transgenes in a process called P-element-dependent silencing (PDS), which is thought to involve the recruitment of heterochromatin proteins. To explore the mechanism of this silencing, we have mobilized KP elements to create a series of strains that contain single, well-defined KP insertions that show PDS. To understand the quantitative role of KP elements in PDS, these single inserts were combined in a series of crosses to obtain genotypes with zero, one, or two KP elements, from which we could examine the effect of KP gene dose. The extent of PDS in these genotypes was shown to be dose dependent in a logarithmic rather than linear fashion. A logarithmic dose dependency is consistent with the KP products interacting with heterochromatic proteins in a concentration-dependent manner such that two molecules are needed to induce gene silencing.

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Heterochromatin Protein 1 Deleted Chromo Domain Decreases Gene Silencing of Transgene in Mouse

Biotechnology Letters ◽

10.1007/s10529-005-6072-4 ◽

2006 ◽

Vol 28 (6) ◽

pp. 419-424 ◽

Cited By ~ 1

Author(s):

Feng-Tao Liu ◽

Yan Zhang

Keyword(s):

Gene Silencing ◽

Heterochromatin Protein 1 ◽

Heterochromatin Protein ◽

Chromo Domain

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Suppression of eNOS-derived superoxide by caveolin-1: a biopterin-dependent mechanism

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00936.2010 ◽

2011 ◽

Vol 301 (3) ◽

pp. H903-H911 ◽

Cited By ~ 33

Author(s):

Kanchana Karuppiah ◽

Lawrence J. Druhan ◽

Chun-an Chen ◽

Travis Smith ◽

Jay L. Zweier ◽

...

Keyword(s):

Gene Silencing ◽

Oxidase Activity ◽

Molecular Mechanisms ◽

Critical Role ◽

Caveolin 1 ◽

Calcium Calmodulin Dependent ◽

Enos Uncoupling ◽

Nadph Oxidation ◽

Interfering Rna

In the vasculature, nitric oxide (NO) is generated by endothelial NO synthase (eNOS) in a calcium/calmodulin-dependent reaction. In the absence of the requisite eNOS cofactor tetrahydrobiopterin (BH4), NADPH oxidation is uncoupled from NO generation, leading to the production of superoxide. Although this phenomenon is apparent with purified enzyme, cellular studies suggest that formation of the BH4 oxidation product, dihydrobiopterin, is the molecular trigger for eNOS uncoupling rather than BH4 depletion alone. In the current study, we investigated the effects of both BH4 depletion and oxidation on eNOS-derived superoxide production in endothelial cells in an attempt to elucidate the molecular mechanisms regulating eNOS oxidase activity. Results demonstrated that pharmacological depletion of endothelial BH4 does not result in eNOS oxidase activity, whereas BH4 oxidation gave rise to significant eNOS-oxidase activity. These findings suggest that the endothelium possesses regulatory mechanisms, which prevent eNOS oxidase activity from pterin-free eNOS. Using a combination of gene silencing and pharmacological approaches, we demonstrate that eNOS-caveolin-1 association is increased under conditions of reduced pterin bioavailability and that this sequestration serves to suppress eNOS uncoupling. Using small interfering RNA approaches, we demonstrate that caveolin-1 gene silencing increases eNOS oxidase activity to 85% of that observed under conditions of BH4 oxidation. Moreover, when caveolin-1 silencing was combined with a pharmacological inhibitor of AKT, BH4 depletion increased eNOS-derived superoxide to 165% of that observed with BH4 oxidation. This study identifies a critical role of caveolin-1 in the regulation of eNOS uncoupling and provides new insight into the mechanisms through which disease-associated changes in caveolin-1 expression may contribute to endothelial dysfunction.

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A critical role of SNF1A/dAMPKα (Drosophila AMP-activated protein kinase α) in muscle on longevity and stress resistance in Drosophila melanogaster

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2010.02.126 ◽

2010 ◽

Vol 394 (1) ◽

pp. 112-118 ◽

Cited By ~ 30

Author(s):

Daisuke Tohyama ◽

Atsushi Yamaguchi

Keyword(s):

Drosophila Melanogaster ◽

Protein Kinase ◽

Stress Resistance ◽

Critical Role ◽

Amp Activated Protein Kinase

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Intramyocellular Fatty-Acid Metabolism Plays a Critical Role in Mediating Responses to Dietary Restriction in Drosophila melanogaster

Cell Metabolism ◽

10.1016/j.cmet.2012.06.005 ◽

2012 ◽

Vol 16 (1) ◽

pp. 97-103 ◽

Cited By ~ 98

Author(s):

Subhash D. Katewa ◽

Fabio Demontis ◽

Marysia Kolipinski ◽

Alan Hubbard ◽

Matthew S. Gill ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Fatty Acid ◽

Fatty Acid Metabolism ◽

Dietary Restriction ◽

Acid Metabolism ◽

Critical Role

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Dual-layer transposon repression in heads of Drosophila melanogaster

10.1101/267039 ◽

2018 ◽

Author(s):

Marius van den Beek ◽

Bruno da Silva ◽

Juliette Pouch ◽

Mohammed el amine Ali Chaouche ◽

Clément Carré ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Gene Silencing ◽

Rna Extraction ◽

Transcript Abundance ◽

Transcriptional Gene Silencing ◽

Loss Of Function ◽

Wild Type ◽

Rna Molecules ◽

Post Transcriptional Gene Silencing ◽

Ping Pong

AbstractpiRNA-mediated repression of transposable elements (TE) in the germline limits the accumulation of heritable mutations caused by their transposition in the genome. It is not clear whether the piRNA pathway plays a functional role in adult, non-gonadal tissues in Drosophila melanogaster. To address this question, we first analyzed the small RNA content of adult Drosophila melanogaster heads. We found that varying amount of piRNA-sized, ping-pong positive molecules in heads correlates with contamination by gonadal tissue during RNA extraction, suggesting that most of piRNAs detected in head sequencing libraries originate from gonads. We next sequenced the heads of wild type and piwi mutants to address whether piwi loss of function would affect the low amount of piRNA-sized, ping-pong negative molecules that are still detected in heads hand-checked to avoid gonadal contamination. We find that loss of piwi does not affect significantly these 24-28 RNA molecules. Instead, we observe increased siRNA levels against the majority of Drosophila transposable element families. To determine the effect of this siRNA level change on transposon expression, we sequenced the transcriptome of wild type, piwi, dicer-2 and piwi, dicer-2 double-mutant fly heads. We find that RNA expression levels of the majority of TE families in piwi or dicer-2 mutants remain unchanged and that TE transcript abundance increases significantly only in piwi, dicer-2 double-mutants. These results lead us to suggest a dual-layer model for TE repression in adult somatic tissues. Piwi-mediated transcriptional gene silencing (TGS) established during embryogenesis constitutes the first layer of TE repression whereas Dicer-2-dependent siRNA-mediated post-transcriptional gene silencing (PTGS) provide a backup mechanism to repress TEs that escape silencing by piwi-mediated TGS.

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Mouse centric and pericentric satellite repeats form distinct functional heterochromatin

The Journal of Cell Biology ◽

10.1083/jcb.200403109 ◽

2004 ◽

Vol 166 (4) ◽

pp. 493-505 ◽

Cited By ~ 309

Author(s):

Mounia Guenatri ◽

Delphine Bailly ◽

Christèle Maison ◽

Geneviève Almouzni

Keyword(s):

Critical Role ◽

Repetitive Sequences ◽

Micrococcal Nuclease ◽

Temporal Isolation ◽

Heterochromatin Protein ◽

Histone Methyltransferases ◽

Centromeric Function ◽

Satellite Repeats ◽

Chromatid Cohesion ◽

Spatio Temporal

Heterochromatin is thought to play a critical role for centromeric function. However, the respective contributions of the distinct repetitive sequences found in these regions, such as minor and major satellites in the mouse, have remained largely unsolved. We show that these centric and pericentric repeats on the chromosomes have distinct heterochromatic characteristics in the nucleus. Major satellites from different chromosomes form clusters associated with heterochromatin protein 1α, whereas minor satellites are individual entities associated with centromeric proteins. Both regions contain methylated histone H3 (Me-K9 H3) but show different micrococcal nuclease sensitivities. A dinucleosome repeating unit is found specifically associated with major satellites. These domains replicate asynchronously, and chromatid cohesion is sustained for a longer time in major satellites compared with minor satellites. Such prolonged cohesion in major satellites is lost in the absence of Suv39h histone methyltransferases. Thus, we define functionally independent centromeric subdomains, which spatio-temporal isolation is proposed to be important for centromeric cohesion and dissociation during chromosome segregation.

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