Raphaëlle Dubruille

Claude Bernard University Lyon 1, Villeurbanne, Rhône-Alpes, France

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Publications (13)109.36 Total impact

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    Benjamin Loppin · Raphaëlle Dubruille · Béatrice Horard
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    ABSTRACT: The union of haploid gametes at fertilization initiates the formation of the diploid zygote in sexually reproducing animals. This founding event of embryogenesis includes several fascinating cellular and nuclear processes, such as sperm-egg cellular interactions, sperm chromatin remodelling, centrosome formation or pronuclear migration. In comparison with other aspects of development, the exploration of animal fertilization at the functional level has remained so far relatively limited, even in classical model organisms. Here, we have reviewed our current knowledge of fertilization in Drosophila melanogaster, with a special emphasis on the genes involved in the complex transformation of the fertilizing sperm nucleus into a replicated set of paternal chromosomes. © 2015 The Authors.
    Full-text · Article · Aug 2015 · Open Biology
  • Raphaëlle Dubruille · Benjamin Loppin
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    ABSTRACT: In Drosophila, telomere capping proteins have the remarkable capacity to recognize chromosome ends in a sequence-independent manner. This epigenetic protection is essential to prevent catastrophic ligations of chromosome extremities. Interestingly, capping proteins occupy a large telomere chromatin domain of several kilobases whose functional relevance to end protection is unknown. Here, we investigated the role of the large capping domain by manipulating HOAP capping protein expression in the male germ cells where telomere protection can be challenged without compromising viability. We show that the exhaustion of HOAP results in a dramatic reduction of other capping proteins at telomeres, including K81, which is essential for male fertility. Strikingly however, we demonstrate that, although capping complexes are barely detected in HOAP-depleted male germ cells, telomere protection and male fertility are not dramatically affected. Our study thus demonstrates that efficient protection of Drosophila telomeres can be achieved with surprisingly low amounts of capping complexes. We propose that these complexes prevent fusions by acting at the very extremity of chromosomes, reminiscent of the protection conferred by extremely short telomeric arrays in yeast or mammalian systems. © 2015. Published by The Company of Biologists Ltd.
    No preview · Article · Apr 2015 · Journal of Cell Science
  • Yong Zhang · Jinli Ling · Chunyan Yuan · Raphaëlle Dubruille · Patrick Emery
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    ABSTRACT: A negative transcriptional feedback loop generates circadian rhythms in Drosophila. PERIOD (PER) is a critical state-variable in this mechanism, and its abundance is tightly regulated. We found that the Drosophila homolog of ATAXIN-2 (ATX2)—an RNA-binding protein implicated in human neurodegenerative diseases—was required for circadian locomotor behavior. ATX2 was necessary for PER accumulation in circadian pacemaker neurons and thus determined period length of circadian behavior. ATX2 was required for the function of TWENTY-FOUR (TYF), a crucial activator of PER translation. ATX2 formed a complex with TYF and promoted its interaction with polyadenylate-binding protein (PABP). Our work uncovers a role for ATX2 in circadian timing and reveals that this protein functions as an activator of PER translation in circadian neurons.
    No preview · Article · May 2013 · Science
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    ABSTRACT: In Drosophila melanogaster, as in many animal and plant species, centromere identity is specified epigenetically. In proliferating cells, a centromere-specific histone H3 variant (CenH3), named Cid in Drosophila and Cenp-A in humans, is a crucial component of the epigenetic centromere mark. Hence, maintenance of the amount and chromosomal location of CenH3 during mitotic proliferation is important. Interestingly, CenH3 may have different roles during meiosis and the onset of embryogenesis. In gametes of Caenorhabditis elegans, and possibly in plants, centromere marking is independent of CenH3. Moreover, male gamete differentiation in animals often includes global nucleosome for protamine exchange that potentially could remove CenH3 nucleosomes. Here we demonstrate that the control of Cid loading during male meiosis is distinct from the regulation observed during the mitotic cycles of early embryogenesis. But Cid is present in mature sperm. After strong Cid depletion in sperm, paternal centromeres fail to integrate into the gonomeric spindle of the first mitosis, resulting in gynogenetic haploid embryos. Furthermore, after moderate depletion, paternal centromeres are unable to re-acquire normal Cid levels in the next generation. We conclude that Cid in sperm is an essential component of the epigenetic centromere mark on paternal chromosomes and it exerts quantitative control over centromeric Cid levels throughout development. Hence, the amount of Cid that is loaded during each cell cycle appears to be determined primarily by the preexisting centromeric Cid, with little flexibility for compensation of accidental losses.
    Full-text · Article · Dec 2012 · PLoS Biology
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    Jinli Ling · Raphaëlle Dubruille · Patrick Emery
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    ABSTRACT: Circadian rhythms are generated by well-conserved interlocked transcriptional feedback loops in animals. In Drosophila, the dimeric transcription factor CLOCK/CYCLE (CLK/CYC) promotes period (per), timeless (tim), vrille (vri), and PAR-domain protein 1 (Pdp1) transcription. PER and TIM negatively feed back on CLK/CYC transcriptional activity, whereas VRI and PDP1 negatively and positively regulate Clk transcription, respectively. Here, we show that the α isoform of the Drosophila FOS homolog KAYAK (KAY) is required for normal circadian behavior. KAY-α downregulation in circadian pacemaker neurons increases period length by 1.5 h. This behavioral phenotype is correlated with decreased expression of several circadian proteins. The strongest effects are on CLK and the neuropeptide PIGMENT DISPERSING FACTOR, which are both under VRI and PDP1 control. Consistently, KAY-α can bind to VRI and inhibit its interaction with the Clk promoter. Interestingly, KAY-α can also repress CLK activity. Hence, in flies with low KAY-α levels, CLK derepression would partially compensate for increased VRI repression, thus attenuating the consequences of KAY-α downregulation on CLK targets. We propose that the double role of KAY-α in the two transcriptional loops controlling Drosophila circadian behavior brings precision and stability to their oscillations.
    Full-text · Article · Nov 2012 · The Journal of Neuroscience : The Official Journal of the Society for Neuroscience
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    Raphaëlle Dubruille · Gabriel A B Marais · Benjamin Loppin
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    ABSTRACT: Comparative genome analysis has allowed the identification of various mechanisms involved in gene birth. However, understanding the evolutionary forces driving new gene origination still represents a major challenge. In particular, an intriguing and not yet fully understood trend has emerged from the study of new genes: many of them show a testis-specific expression pattern, which has remained poorly understood. Here we review the case of such a new gene, which involves a telomere-capping gene family in Drosophila. hiphop and its testis-specific paralog K81 are critical for the protection of chromosome ends in somatic cells and male gametes, respectively. Two independent functional studies recently proposed that these genes evolved under a reproductive-subfunctionalization regime. The 2011 release of new Drosophila genome sequences from the melanogaster group of species allowed us to deepen our phylogenetic analysis of the hiphop/K81 family. This work reveals an unsuspected dynamic of gene birth and death within the group, with recurrent duplication events through retroposition mechanisms. Finally, we discuss the plausibility of different evolutionary scenarios that could explain the diversification of this gene family.
    Preview · Article · Jul 2012
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    Raphaëlle Dubruille · Benjamin Loppin
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    ABSTRACT: A critical function of telomeres is to prevent the ligation of chromosome ends by DNA repair enzymes. In most eukaryotes, telomeric DNA consists in large arrays of G-rich tandem repeats that are recognized by DNA binding capping proteins. Drosophila telomeres are unusual as they lack short tandem repeats. However, Drosophila capping proteins can bind chromosome extremities in a DNA sequence-independent manner. This epigenetic protection of fly telomeres has been essentially studied in somatic cells where capping proteins such as HOAP or HP1 are essential in preventing chromosome end-to-end fusions. HipHop and K81 are two recently identified paralogous capping proteins with complementary expression patterns. While HipHop is involved in telomere capping in somatic cells, K81 has specialized in the protection of telomeres in post-meiotic male germ cells. Remarkably, K81 is required for the stabilization of HOAP and HP1 at telomeres during the massive paternal chromatin remodeling that occurs during spermiogenesis and at fertilization. We thus propose that the maintenance of capping proteins at Drosophila sperm telomeres is crucial for the transmission of telomere identity to the diploid zygote. :
    Preview · Article · Apr 2011 · Cell cycle (Georgetown, Tex.)
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    ABSTRACT: A critical function of telomeres is to prevent fusion of chromosome ends by the DNA repair machinery. In Drosophila somatic cells, assembly of the protecting capping complex at telomeres notably involves the recruitment of HOAP, HP1, and their recently identified partner, HipHop. We previously showed that the hiphop gene was duplicated before the radiation of the melanogaster subgroup of species, giving birth to K81, a unique paternal effect gene specifically expressed in the male germline. Here we show that K81 specifically associates with telomeres during spermiogenesis, along with HOAP and HP1, and is retained on paternal chromosomes until zygote formation. In K81 mutant testes, capping proteins are not maintained at telomeres in differentiating spermatids, resulting in the transmission of uncapped paternal chromosomes that fail to properly divide during the first zygotic mitosis. Despite the apparent similar capping roles of K81 and HipHop in their respective domain of expression, we demonstrate by in vivo reciprocal complementation analyses that they are not interchangeable. Strikingly, HipHop appeared to be unable to maintain capping proteins at telomeres during the global chromatin remodeling of spermatid nuclei. Our data demonstrate that K81 is essential for the maintenance of capping proteins at telomeres in postmeiotic male germ cells. In species of the melanogaster subgroup, HipHop and K81 have not only acquired complementary expression domains, they have also functionally diverged following the gene duplication event. We propose that K81 specialized in the maintenance of telomere protection in the highly peculiar chromatin environment of differentiating male gametes.
    Full-text · Article · Dec 2010 · Current biology: CB
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    Dataset: Figure S3
    Raphaëlle Dubruille · Alejandro Murad · Michael Rosbash · Patrick Emery
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    ABSTRACT: cry overexpression restores constant light arrhythmicity in flies expressing kis dsRNAs. (A) Actograms showing the locomotor activity of tim-GAL4 VDRC46685/+ (tg4 VDRC46685;+); and tim-GAL4 VDRC46685/UAS-cry flies (tg4 VDRC46685;cry) under LL conditions. Both genotypes are homozygous for ls-tim (n = 16 flies for each genotype). (B) Percentage of LL rhythmicity for the same genotypes. (1.32 MB TIF)
    Preview · Dataset · Dec 2009
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    Dataset: Figure S2
    Raphaëlle Dubruille · Alejandro Murad · Michael Rosbash · Patrick Emery
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    ABSTRACT: KIS protein levels are severely reduced in circadian neurons expressing kis dsRNAs. (A) KIS protein levels in large and small LNvs and in LNds measured by immunostaining. The histogram represents the normalized KIS fluorescence signal (see Materials and Methods for details), measured in control flies (pdf-GAL4 UAS-dcr2/+ [pg4 dcr2/+] and tim-GAL4 UAS-dcr2; R32 [tg4 dcr2/+]) and flies expressing kis dsRNAs in all circadian neurons (tim-GAL4 UAS-dcr2/VDRC46685; R32 [tg4 dcr2/VDRC46685]) or only in PDF positive neurons (pdf-GAL4 UAS-dcr2/VDRC46685 [pg4 dcr2/VDRC46685]). KIS expression is reduced by about 85–90% in both mutant genotypes, and in all cell types surveyed. When driving kis dsRNAs with pdf-GAL4, we used an anti-PDF antibody to identify the PDF positive LNvs. With tim-GAL4, we used flies carrying one copy of the R32 lacZ insertion trap [58] and identified clock neurons with an anti-βGAL antibody. βGAL staining was weak in Dorsal Neurons, but a few DN1s could nevertheless be identified in tg4 dcr2/VDRC46685 brains. KIS expression was also severely reduced in these neurons (data not shown). Error bars represent standard deviations. (B) Histogram showing the percentage of rhythmicity in constant light for control flies (pdf-GAL4 UAS-dcr2/+ [pg4 dcr2/+] and tim-GAL4 UAS-dcr2/+ [tg4 dcr2/+]) and flies expressing kis dsRNAs in all circadian neurons (tim-GAL4 UAS-dcr2/VDRC46685 [tg4 dcr2/VDRC46685]; ls-tim homozygotes) or only in PDF positive neurons (pdf-GAL4 UAS-dcr2/VDRC46685 [pg4 dcr2/VDRC46685]; ls-tim homozygotes) (n = 16 flies for each genotype). As also shown on Figure 5 with the NIG-Fly line, driving the VDRC kis dsRNAs only in PDF positive LNvs does not induce LL rhythmicity, even though as shown in (A) KIS expression is as efficiently repressed as with tim-GAL4. AR: complete arrhythmicity. (0.51 MB TIF)
    Preview · Dataset · Dec 2009
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    Dataset: Table S2
    Raphaëlle Dubruille · Alejandro Murad · Michael Rosbash · Patrick Emery
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    ABSTRACT: Behavior of EP(2)2356 crossed to different GAL4 drivers and GAL80 repressor transgenes under constant light (200 lux, unless otherwise indicted) (AR = arrhythmicity). (0.03 MB DOC)
    Preview · Dataset · Dec 2009
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    Dataset: Table S1
    Raphaëlle Dubruille · Alejandro Murad · Michael Rosbash · Patrick Emery
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    ABSTRACT: Behavior of the selected EP lines crossed to tim-GAL4 under constant darkness (as = antisense orientation). (0.07 MB DOC)
    Preview · Dataset · Dec 2009
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    Raphaëlle Dubruille · Alejandro D. Murad · Michael Rosbash · Patrick Emery
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    ABSTRACT: Circadian pacemakers are essential to synchronize animal physiology and behavior with the dayrationight cycle. They are self-sustained, but the phase of their oscillations is determined by environmental cues, particularly light intensity and temperature cycles. In Drosophila, light is primarily detected by a dedicated blue-light photoreceptor: CRYPTOCHROME (CRY). Upon light activation, CRY binds to the pacemaker protein TIMELESS (TIM) and triggers its proteasomal degradation, thus resetting the circadian pacemaker. To understand further the CRY input pathway, we conducted a misexpression screen under constant light based on the observation that flies with a disruption in the CRY input pathway remain robustly rhythmic instead of becoming behaviorally arrhythmic. We report the identification of more than 20 potential regulators of CRY-dependent light responses. We demonstrate that one of them, the chromatin-remodeling enzyme KISMET (KIS), is necessary for normal circadian photoresponses, but does not affect the circadian pacemaker. KIS genetically interacts with CRY and functions in PDF-negative circadian neurons, which play an important role in circadian light responses. It also affects daily CRY-dependent TIM oscillations in a peripheral tissue: the eyes. We therefore conclude that KIS is a key transcriptional regulator of genes that function in the CRY signaling cascade, and thus it plays an important role in the synchronization of circadian rhythms with the dayrationight cycle.
    Full-text · Article · Dec 2009 · PLoS Genetics
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    Raphaelle Dubruille · Patrick Emery
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    ABSTRACT: Circadian clocks synchronize the physiology and behavior of most animals with the day to night cycle. A fundamental property of the molecular pacemakers generating circadian rhythms is their self-sustained nature: they keep oscillating even under constant conditions, with a period close to, but not exactly, 24 h. However, circadian pacemakers have to be sensitive to environmental cues to be beneficial. They need to be reset every day to keep a proper phase relationship with the day to night cycle, and they have to be able to adjust to seasonal changes in day length and temperature. Here, we review our current knowledge of the molecular and neural mechanisms contributing to the plasticity of Drosophila circadian rhythms, which are proving to be remarkably sophisticated and complex.
    Full-text · Article · Sep 2008 · Molecular Neurobiology
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    ABSTRACT: Regulatory factor X (RFX) transcription factors play a key role in ciliary assembly in nematode, Drosophila and mouse. Using the tremendous advantages of comparative genomics in closely related species, we identified novel genes regulated by dRFX in Drosophila. We first demonstrate that a subset of known ciliary genes in Caenorhabditis elegans and Drosophila are regulated by dRFX and have a conserved RFX binding site (X-box) in their promoters in two highly divergent Drosophila species. We then designed an X-box consensus sequence and carried out a genome wide computer screen to identify novel genes under RFX control. We found 412 genes that share a conserved X-box upstream of the ATG in both species, with 83 genes presenting a more restricted consensus. We analyzed 25 of these 83 genes, 16 of which are indeed RFX target genes. Two of them have never been described as involved in ciliogenesis. In addition, reporter construct expression analysis revealed that three of the identified genes encode proteins specifically localized in ciliated endings of Drosophila sensory neurons. Our X-box search strategy led to the identification of novel RFX target genes in Drosophila that are involved in sensory ciliogenesis. We also established a highly valuable Drosophila cilia and basal body dataset. These results demonstrate the accuracy of the X-box screen and will be useful for the identification of candidate genes for human ciliopathies, as several human homologs of RFX target genes are known to be involved in diseases, such as Bardet-Biedl syndrome.
    Full-text · Article · Feb 2007 · Genome biology
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    ABSTRACT: There are five members of the RFX family of transcription factors in mammals. While RFX5 plays a well-defined role in the immune system, the functions of RFX1 to RFX4 remain largely unknown. We have generated mice with a deletion of the Rfx3 gene. RFX3-deficient mice exhibit frequent left-right (LR) asymmetry defects leading to a high rate of embryonic lethality and situs inversus in surviving adults. In vertebrates, specification of the LR body axis is controlled by monocilia in the embryonic node, and defects in nodal cilia consequently result in abnormal LR patterning. Consistent with this, Rfx3 is expressed in ciliated cells of the node and RFX3-deficient mice exhibit a pronounced defect in nodal cilia. In contrast to the case for wild-type embryos, for which we document for the first time a twofold increase in the length of nodal cilia during development, the cilia are present but remain markedly stunted in mutant embryos. Finally, we show that RFX3 regulates the expression of D2lic, the mouse orthologue of a Caenorhabditis elegans gene that is implicated in intraflagellar transport, a process required for the assembly and maintenance of cilia. In conclusion, RFX3 is essential for the differentiation of nodal monocilia and hence for LR body axis determination.
    Full-text · Article · Jun 2004 · Molecular and Cellular Biology
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    ABSTRACT: Ciliated neurons play an important role in sensory perception in many animals. Modified cilia at dendrite endings serve as sites of sensory signal capture and transduction. We describe Drosophila mutations that affect the transcription factor RFX and genetic rescue experiments that demonstrate its central role in sensory cilium differentiation. Rfx mutant flies show defects in chemosensory and mechanosensory behaviors but have normal phototaxis, consistent with Rfx expression in ciliated sensory neurons and neuronal precursors but not in photoreceptors. The mutant behavioral phenotypes are correlated with abnormal function and structure of neuronal cilia, as shown by the loss of sensory transduction and by defects in ciliary morphology and ultrastructure. These results identify Rfx as an essential regulator of ciliated sensory neuron differentiation in Drosophila.
    No preview · Article · Jan 2003 · Development

Publication Stats

465 Citations
109.36 Total Impact Points

Institutions

  • 2004-2015
    • Claude Bernard University Lyon 1
      • Centre de génétique et de physiologie moléculaire et cellulaire (CGphyMC)
      Villeurbanne, Rhône-Alpes, France
  • 2007-2013
    • University of Massachusetts Medical School
      • Department of Neurobiology
      Worcester, Massachusetts, United States
  • 2003-2012
    • French National Centre for Scientific Research
      • Centre of Molecular Genetics
      Lutetia Parisorum, Île-de-France, France