The Experts below are selected from a list of 174 Experts worldwide ranked by ideXlab platform
Mireille Bétermier - One of the best experts on this subject based on the ideXlab platform.
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RESEARCH ARTICLE TFIIS-Dependent Non-coding Transcription Regulates Developmental Genome Rearrangements
2016Co-Authors: Kamila Maliszewska-olejniczak, Nathalie Mathy, Olivier Arnaiz, Julita Gruchota, Robert Gromadka, Cyril Denby, Ra Duharcourt, Mireille BétermierAbstract:Because of their Nuclear Dimorphism, ciliates provide a unique opportunity to study the role of non-coding RNAs (ncRNAs) in the communication between germline and somatic line-ages. In these unicellular eukaryotes, a new somatic nucleus develops at each sexual cycle from a copy of the zygotic (germline) nucleus, while the old somatic nucleus degenerates. In the ciliate Paramecium tetraurelia, the genome is massively rearranged during this pro-cess through the reproducible elimination of repeated sequences and the precise excision of over 45,000 short, single-copy Internal Eliminated Sequences (IESs). Different types of ncRNAs resulting from genome-wide transcription were shown to be involved in the epige-netic regulation of genome rearrangements. To understand how ncRNAs are produced from the entire genome, we have focused on a homolog of the TFIIS elongation factor, which regulates RNA polymerase II transcriptional pausing. Six TFIIS-paralogs, represent-ing four distinct families, can be found in P. tetraurelia genome. Using RNA interference, we showed that TFIIS4, which encodes a development-specific TFIIS protein, is essential for the formation of a functional somatic genome. Molecular analyses and high-throughpu
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TFIIS-Dependent Non-coding Transcription Regulates Developmental Genome Rearrangements.
PLoS Genetics, 2015Co-Authors: Kamila Maliszewska-olejniczak, Nathalie Mathy, Olivier Arnaiz, Julita Gruchota, Robert Gromadka, Mireille Bétermier, Sandra Duharcourt, Cyril Denby Wilkes, Jacek K NowakAbstract:Because of their Nuclear Dimorphism, ciliates provide a unique opportunity to study the role of non-coding RNAs (ncRNAs) in the communication between germline and somatic lineages. In these unicellular eukaryotes, a new somatic nucleus develops at each sexual cycle from a copy of the zygotic (germline) nucleus, while the old somatic nucleus degenerates. In the ciliate Paramecium tetraurelia, the genome is massively rearranged during this process through the reproducible elimination of repeated sequences and the precise excision of over 45,000 short, single-copy Internal Eliminated Sequences (IESs). Different types of ncRNAs resulting from genome-wide transcription were shown to be involved in the epigenetic regulation of genome rearrangements. To understand how ncRNAs are produced from the entire genome, we have focused on a homolog of the TFIIS elongation factor, which regulates RNA polymerase II transcriptional pausing. Six TFIIS-paralogs, representing four distinct families, can be found in P. tetraurelia genome. Using RNA interference, we showed that TFIIS4, which encodes a development-specific TFIIS protein, is essential for the formation of a functional somatic genome. Molecular analyses and high-throughput DNA sequencing upon TFIIS4 RNAi demonstrated that TFIIS4 is involved in all kinds of genome rearrangements, including excision of ~48% of IESs. Localization of a GFP-TFIIS4 fusion revealed that TFIIS4 appears specifically in the new somatic nucleus at an early developmental stage, before IES excision. RT-PCR experiments showed that TFIIS4 is necessary for the synthesis of IES-containing non-coding transcripts. We propose that these IES+ transcripts originate from the developing somatic nucleus and serve as pairing substrates for germline-specific short RNAs that target elimination of their homologous sequences. Our study, therefore, connects the onset of zygotic non coding transcription to the control of genome plasticity in Paramecium, and establishes for the first time a specific role of TFIIS in non-coding transcription in eukaryotes.
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ku mediated coupling of dna cleavage and repair during programmed genome rearrangements in the ciliate paramecium tetraurelia
PLOS Genetics, 2014Co-Authors: Antoine Marmignon, Sophie Malinsky, Olivier Arnaiz, Emeline Dubois, Julien Bischerour, Aude Silve, Clementine Fojcik, Aurelie Kapusta, Mireille BétermierAbstract:During somatic differentiation, physiological DNA double-strand breaks (DSB) can drive programmed genome rearrangements (PGR), during which DSB repair pathways are mobilized to safeguard genome integrity. Because of their unique Nuclear Dimorphism, ciliates are powerful unicellular eukaryotic models to study the mechanisms involved in PGR. At each sexual cycle, the germline nucleus is transmitted to the progeny, but the somatic nucleus, essential for gene expression, is destroyed and a new somatic nucleus differentiates from a copy of the germline nucleus. In Paramecium tetraurelia, the development of the somatic nucleus involves massive PGR, including the precise elimination of at least 45,000 germline sequences (Internal Eliminated Sequences, IES). IES excision proceeds through a cut-and-close mechanism: a domesticated transposase, PiggyMac, is essential for DNA cleavage, and DSB repair at excision sites involves the Ligase IV, a specific component of the non-homologous end-joining (NHEJ) pathway. At the genome-wide level, a huge number of programmed DSBs must be repaired during this process to allow the assembly of functional somatic chromosomes. To understand how DNA cleavage and DSB repair are coordinated during PGR, we have focused on Ku, the earliest actor of NHEJ-mediated repair. Two Ku70 and three Ku80 paralogs are encoded in the genome of P. tetraurelia: Ku70a and Ku80c are produced during sexual processes and localize specifically in the developing new somatic nucleus. Using RNA interference, we show that the development-specific Ku70/Ku80c heterodimer is essential for the recovery of a functional somatic nucleus. Strikingly, at the molecular level, PiggyMac-dependent DNA cleavage is abolished at IES boundaries in cells depleted for Ku80c, resulting in IES retention in the somatic genome. PiggyMac and Ku70a/Ku80c co-purify as a complex when overproduced in a heterologous system. We conclude that Ku has been integrated in the Paramecium DNA cleavage factory, enabling tight coupling between DSB introduction and repair during PGR.
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transposon invasion of the paramecium germline genome countered by a domesticated piggybac transposase and the nhej pathway
International Journal of Evolutionary Biology, 2012Co-Authors: Nathalie Mathy, Emeline Dubois, Julien Bischerour, Antoine Marmignon, Vinciane Regnier, Mireille BétermierAbstract:Sequences related to transposons constitute a large fraction of extant genomes, but insertions within coding sequences have generally not been tolerated during evolution. Thanks to their unique Nuclear Dimorphism and to their original mechanism of programmed DNA elimination from their somatic nucleus (macronucleus), ciliates are emerging model organisms for the study of the impact of transposable elements on genomes. The germline genome of the ciliate Paramecium, located in its micronucleus, contains thousands of short intervening sequences, the IESs, which interrupt 47% of genes. Recent data provided support to the hypothesis that an evolutionary link exists between Paramecium IESs and Tc1/mariner transposons. During development of the macronucleus, IESs are excised precisely thanks to the coordinated action of PiggyMac, a domesticated piggyBac transposase, and of the NHEJ double-strand break repair pathway. A PiggyMac homolog is also required for developmentally programmed DNA elimination in another ciliate, Tetrahymena. Here, we present an overview of the life cycle of these unicellular eukaryotes and of the developmentally programmed genome rearrangements that take place at each sexual cycle. We discuss how ancient domestication of a piggyBac transposase might have allowed Tc1/mariner elements to spread throughout the germline genome of Paramecium, without strong counterselection against insertion within genes.
Olivier Arnaiz - One of the best experts on this subject based on the ideXlab platform.
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RESEARCH ARTICLE TFIIS-Dependent Non-coding Transcription Regulates Developmental Genome Rearrangements
2016Co-Authors: Kamila Maliszewska-olejniczak, Nathalie Mathy, Olivier Arnaiz, Julita Gruchota, Robert Gromadka, Cyril Denby, Ra Duharcourt, Mireille BétermierAbstract:Because of their Nuclear Dimorphism, ciliates provide a unique opportunity to study the role of non-coding RNAs (ncRNAs) in the communication between germline and somatic line-ages. In these unicellular eukaryotes, a new somatic nucleus develops at each sexual cycle from a copy of the zygotic (germline) nucleus, while the old somatic nucleus degenerates. In the ciliate Paramecium tetraurelia, the genome is massively rearranged during this pro-cess through the reproducible elimination of repeated sequences and the precise excision of over 45,000 short, single-copy Internal Eliminated Sequences (IESs). Different types of ncRNAs resulting from genome-wide transcription were shown to be involved in the epige-netic regulation of genome rearrangements. To understand how ncRNAs are produced from the entire genome, we have focused on a homolog of the TFIIS elongation factor, which regulates RNA polymerase II transcriptional pausing. Six TFIIS-paralogs, represent-ing four distinct families, can be found in P. tetraurelia genome. Using RNA interference, we showed that TFIIS4, which encodes a development-specific TFIIS protein, is essential for the formation of a functional somatic genome. Molecular analyses and high-throughpu
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Ku-Mediated Coupling of DNA Cleavage and Repair during Programmed Genome Rearrangements in the
2016Co-Authors: Ciliate Paramecium Tetraurelia, Olivier Arnaiz, Emeline Dubois, Julien Bischerour, Antoine Marmignon, Aude Silve, Sophie MalinskyAbstract:During somatic differentiation, physiological DNA double-strand breaks (DSB) can drive programmed genome rearrangements (PGR), during which DSB repair pathways are mobilized to safeguard genome integrity. Because of their unique Nuclear Dimorphism, ciliates are powerful unicellular eukaryotic models to study the mechanisms involved in PGR. At each sexual cycle, the germline nucleus is transmitted to the progeny, but the somatic nucleus, essential for gene expression, is destroyed and a new somatic nucleus differentiates from a copy of the germline nucleus. In Paramecium tetraurelia, the development of the somatic nucleus involves massive PGR, including the precise elimination of at least 45,000 germline sequences (Internal Eliminated Sequences, IES). IES excision proceeds through a cut-and-close mechanism: a domesticated transposase, PiggyMac, is essential for DNA cleavage, and DSB repair at excision sites involves the Ligase IV, a specific component of the non-homologous end-joining (NHEJ) pathway. At the genome-wide level, a huge number of programmed DSBs must be repaired during this process to allow the assembly of functional somatic chromosomes. To understand how DNA cleavage and DSB repair are coordinated during PGR, we have focused on Ku, the earliest actor of NHEJ-mediated repair. Two Ku70 and three Ku80 paralogs are encoded in the genome of P. tetraurelia: Ku70a and Ku80c are produced during sexual processes and localize specifically in the developing new somatic nucleus. Using RNA interference, we show that the development-specific Ku70/Ku80c heterodimer is essential for the recovery of a functional somati
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TFIIS-Dependent Non-coding Transcription Regulates Developmental Genome Rearrangements.
PLoS Genetics, 2015Co-Authors: Kamila Maliszewska-olejniczak, Nathalie Mathy, Olivier Arnaiz, Julita Gruchota, Robert Gromadka, Mireille Bétermier, Sandra Duharcourt, Cyril Denby Wilkes, Jacek K NowakAbstract:Because of their Nuclear Dimorphism, ciliates provide a unique opportunity to study the role of non-coding RNAs (ncRNAs) in the communication between germline and somatic lineages. In these unicellular eukaryotes, a new somatic nucleus develops at each sexual cycle from a copy of the zygotic (germline) nucleus, while the old somatic nucleus degenerates. In the ciliate Paramecium tetraurelia, the genome is massively rearranged during this process through the reproducible elimination of repeated sequences and the precise excision of over 45,000 short, single-copy Internal Eliminated Sequences (IESs). Different types of ncRNAs resulting from genome-wide transcription were shown to be involved in the epigenetic regulation of genome rearrangements. To understand how ncRNAs are produced from the entire genome, we have focused on a homolog of the TFIIS elongation factor, which regulates RNA polymerase II transcriptional pausing. Six TFIIS-paralogs, representing four distinct families, can be found in P. tetraurelia genome. Using RNA interference, we showed that TFIIS4, which encodes a development-specific TFIIS protein, is essential for the formation of a functional somatic genome. Molecular analyses and high-throughput DNA sequencing upon TFIIS4 RNAi demonstrated that TFIIS4 is involved in all kinds of genome rearrangements, including excision of ~48% of IESs. Localization of a GFP-TFIIS4 fusion revealed that TFIIS4 appears specifically in the new somatic nucleus at an early developmental stage, before IES excision. RT-PCR experiments showed that TFIIS4 is necessary for the synthesis of IES-containing non-coding transcripts. We propose that these IES+ transcripts originate from the developing somatic nucleus and serve as pairing substrates for germline-specific short RNAs that target elimination of their homologous sequences. Our study, therefore, connects the onset of zygotic non coding transcription to the control of genome plasticity in Paramecium, and establishes for the first time a specific role of TFIIS in non-coding transcription in eukaryotes.
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ku mediated coupling of dna cleavage and repair during programmed genome rearrangements in the ciliate paramecium tetraurelia
PLOS Genetics, 2014Co-Authors: Antoine Marmignon, Sophie Malinsky, Olivier Arnaiz, Emeline Dubois, Julien Bischerour, Aude Silve, Clementine Fojcik, Aurelie Kapusta, Mireille BétermierAbstract:During somatic differentiation, physiological DNA double-strand breaks (DSB) can drive programmed genome rearrangements (PGR), during which DSB repair pathways are mobilized to safeguard genome integrity. Because of their unique Nuclear Dimorphism, ciliates are powerful unicellular eukaryotic models to study the mechanisms involved in PGR. At each sexual cycle, the germline nucleus is transmitted to the progeny, but the somatic nucleus, essential for gene expression, is destroyed and a new somatic nucleus differentiates from a copy of the germline nucleus. In Paramecium tetraurelia, the development of the somatic nucleus involves massive PGR, including the precise elimination of at least 45,000 germline sequences (Internal Eliminated Sequences, IES). IES excision proceeds through a cut-and-close mechanism: a domesticated transposase, PiggyMac, is essential for DNA cleavage, and DSB repair at excision sites involves the Ligase IV, a specific component of the non-homologous end-joining (NHEJ) pathway. At the genome-wide level, a huge number of programmed DSBs must be repaired during this process to allow the assembly of functional somatic chromosomes. To understand how DNA cleavage and DSB repair are coordinated during PGR, we have focused on Ku, the earliest actor of NHEJ-mediated repair. Two Ku70 and three Ku80 paralogs are encoded in the genome of P. tetraurelia: Ku70a and Ku80c are produced during sexual processes and localize specifically in the developing new somatic nucleus. Using RNA interference, we show that the development-specific Ku70/Ku80c heterodimer is essential for the recovery of a functional somatic nucleus. Strikingly, at the molecular level, PiggyMac-dependent DNA cleavage is abolished at IES boundaries in cells depleted for Ku80c, resulting in IES retention in the somatic genome. PiggyMac and Ku70a/Ku80c co-purify as a complex when overproduced in a heterologous system. We conclude that Ku has been integrated in the Paramecium DNA cleavage factory, enabling tight coupling between DSB introduction and repair during PGR.
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The Paramecium Germline Genome Provides a Niche for Intragenic Parasitic DNA: Evolutionary Dynamics of Internal Eliminated Sequences
PLOS Genetics, 2012Co-Authors: Olivier Arnaiz, Nathalie Mathy, Céline Baudry, Sophie Malinsky, Cyril Denby Wilkes, Benjamin E. Lauderdale, Jean-marc Aury, Olivier Garnier, Karine Labadie, Anne Le MouëlAbstract:Insertions of parasitic DNA within coding sequences are usually deleterious and are generally counter-selected during evolution. Thanks to Nuclear Dimorphism, ciliates provide unique models to study the fate of such insertions. Their germline genome undergoes extensive rearrangements during development of a new somatic macronucleus from the germline micronucleus following sexual events. In Paramecium, these rearrangements include precise excision of unique-copy Internal Eliminated Sequences (IES) from the somatic DNA, requiring the activity of a domesticated piggyBac transposase, PiggyMac. We have sequenced Paramecium tetraurelia germline DNA, establishing a genome-wide catalogue of ∼45,000 IESs, in order to gain insight into their evolutionary origin and excision mechanism. We obtained direct evidence that PiggyMac is required for excision of all IESs. Homology with known P. tetraurelia Tc1/mariner transposons, described here, indicates that at least a fraction of IESs derive from these elements. Most IES insertions occurred before a recent whole-genome duplication that preceded diversification of the P. aurelia species complex, but IES invasion of the Paramecium genome appears to be an ongoing process. Once inserted, IESs decay rapidly by accumulation of deletions and point substitutions. Over 90% of the IESs are shorter than 150 bp and present a remarkable size distribution with a ∼10 bp periodicity, corresponding to the helical repeat of double-stranded DNA and suggesting DNA loop formation during assembly of a transpososome-like excision complex. IESs are equally frequent within and between coding sequences; however, excision is not 100% efficient and there is selective pressure against IES insertions, in particular within highly expressed genes. We discuss the possibility that ancient domestication of a piggyBac transposase favored subsequent propagation of transposons throughout the germline by allowing insertions in coding sequences, a fraction of the genome in which parasitic DNA is not usually tolerated.
Laura F. Landweber - One of the best experts on this subject based on the ideXlab platform.
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Transcribed germline-limited coding sequences in Oxytricha trifallax.
G3 (Bethesda Md.), 2021Co-Authors: Richard V Miller, V Talya Yerlici, Rafik Neme, Derek M Clay, Jananan S Pathmanathan, Jaspreet S Khurana, Laura F. LandweberAbstract:The germline-soma divide is a fundamental distinction in developmental biology, and different genes are expressed in germline and somatic cells throughout metazoan life cycles. Ciliates, a group of microbial eukaryotes, exhibit germline-somatic Nuclear Dimorphism within a single cell with two different genomes. The ciliate Oxytricha trifallax undergoes massive RNA-guided DNA elimination and genome rearrangement to produce a new somatic macronucleus (MAC) from a copy of the germline micronucleus (MIC). This process eliminates noncoding DNA sequences that interrupt genes and also deletes hundreds of germline-limited open reading frames (ORFs) that are transcribed during genome rearrangement. Here, we update the set of transcribed germline-limited ORFs (TGLOs) in O. trifallax. We show that TGLOs tend to be expressed during Nuclear development and then are absent from the somatic MAC. We also demonstrate that exposure to synthetic RNA can reprogram TGLO retention in the somatic MAC and that TGLO retention leads to transcription outside the normal developmental program. These data suggest that TGLOs represent a group of developmentally regulated protein coding sequences whose gene expression is terminated by DNA elimination.
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a database of ciliate genome rearrangements
Nucleic Acids Research, 2015Co-Authors: Jonathan Burns, Denys Kukushkin, Kelsi A. Lindblad, Xiao Chen, Nataša Jonoska, Laura F. LandweberAbstract:Ciliated protists exhibit Nuclear Dimorphism through the presence of somatic macronuclei (MAC) and germline micronuclei (MIC). In some ciliates, DNA from precursor segments in the MIC genome rearranges to form transcriptionally active genes in the mature MAC genome, making these ciliates model organisms to study the process of somatic genome rearrangement. Similar broad scale, somatic rearrangement events occur in many eukaryotic cells and tumors. The (http://oxytricha.princeton.edu/mds_ies_db) is a database of genome recombination and rearrangement annotations, and it provides tools for visualization and comparative analysis of precursor and product genomes. The database currently contains annotations for two completely sequenced ciliate genomes: Oxytricha trifallax and Tetrahymena thermophila.
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Programmed Genome Rearrangements in the Ciliate Oxytricha.
Microbiology spectrum, 2015Co-Authors: V Talya Yerlici, Laura F. LandweberAbstract:The ciliate Oxytricha is a microbial eukaryote with two genomes, one of which experiences extensive genome remodeling during development. Each round of conjugation initiates a cascade of events that construct a transcriptionally active somatic genome from a scrambled germline genome, with considerable help from both long and small noncoding RNAs. This process of genome remodeling entails massive DNA deletion and reshuffling of remaining DNA segments to form functional genes from their interrupted and scrambled germline precursors. The use of Oxytricha as a model system provides an opportunity to study an exaggerated form of programmed genome rearrangement. Furthermore, studying the mechanisms that maintain Nuclear Dimorphism and mediate genome rearrangement has demonstrated a surprising plasticity and diversity of noncoding RNA pathways, with new roles that go beyond conventional gene silencing. Another aspect of ciliate genetics is their unorthodox patterns of RNA-mediated, epigenetic inheritance that rival Mendelian inheritance. This review takes the reader through the key experiments in a model eukaryote that led to fundamental discoveries in RNA biology and pushes the biological limits of DNA processing.
F. Paul Doerder - One of the best experts on this subject based on the ideXlab platform.
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The “Tetrahymena pyriformis” complex of cryptic species
Biodiversity and Conservation, 2008Co-Authors: Ellen M. Simon, David L. Nanney, F. Paul DoerderAbstract:Cryptic species are common among protists and have long been known in ciliates. The ciliate genus Tetrahymena contains a large group of morphologically indistinguishable species referred to as the ‘ T. pyriformis ’ complex. These species include those reproductively isolated by mating type as well as asexual species characterized by the absence of the germinal micronucleus. This paper examines the molecular diversity of the species and describes the biogeography of ‘ T. pyriformis ’ species. Most species are globally distributed, though the best studied species, T. thermophila , is confined to North America and gives evidence of population structure in local populations. Selfers and asexual species are common and arise from sexual species, a possible exploitation of Nuclear Dimorphism. It is argued that the cryptic species likely have different ecological roles and that the biodiversity of Tetrahymena in particular, and ciliates in general, is underestimated.
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The “ Tetrahymena pyriformis ” complex of cryptic species
Biodiversity and Conservation, 2007Co-Authors: Ellen M. Simon, David L. Nanney, F. Paul DoerderAbstract:Cryptic species are common among protists and have long been known in ciliates. The ciliate genus Tetrahymena contains a large group of morphologically indistinguishable species referred to as the ‘T. pyriformis’ complex. These species include those reproductively isolated by mating type as well as asexual species characterized by the absence of the germinal micronucleus. This paper examines the molecular diversity of the species and describes the biogeography of ‘T. pyriformis’ species. Most species are globally distributed, though the best studied species, T. thermophila, is confined to North America and gives evidence of population structure in local populations. Selfers and asexual species are common and arise from sexual species, a possible exploitation of Nuclear Dimorphism. It is argued that the cryptic species likely have different ecological roles and that the biodiversity of Tetrahymena in particular, and ciliates in general, is underestimated.
Nathalie Mathy - One of the best experts on this subject based on the ideXlab platform.
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RESEARCH ARTICLE TFIIS-Dependent Non-coding Transcription Regulates Developmental Genome Rearrangements
2016Co-Authors: Kamila Maliszewska-olejniczak, Nathalie Mathy, Olivier Arnaiz, Julita Gruchota, Robert Gromadka, Cyril Denby, Ra Duharcourt, Mireille BétermierAbstract:Because of their Nuclear Dimorphism, ciliates provide a unique opportunity to study the role of non-coding RNAs (ncRNAs) in the communication between germline and somatic line-ages. In these unicellular eukaryotes, a new somatic nucleus develops at each sexual cycle from a copy of the zygotic (germline) nucleus, while the old somatic nucleus degenerates. In the ciliate Paramecium tetraurelia, the genome is massively rearranged during this pro-cess through the reproducible elimination of repeated sequences and the precise excision of over 45,000 short, single-copy Internal Eliminated Sequences (IESs). Different types of ncRNAs resulting from genome-wide transcription were shown to be involved in the epige-netic regulation of genome rearrangements. To understand how ncRNAs are produced from the entire genome, we have focused on a homolog of the TFIIS elongation factor, which regulates RNA polymerase II transcriptional pausing. Six TFIIS-paralogs, represent-ing four distinct families, can be found in P. tetraurelia genome. Using RNA interference, we showed that TFIIS4, which encodes a development-specific TFIIS protein, is essential for the formation of a functional somatic genome. Molecular analyses and high-throughpu
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TFIIS-Dependent Non-coding Transcription Regulates Developmental Genome Rearrangements.
PLoS Genetics, 2015Co-Authors: Kamila Maliszewska-olejniczak, Nathalie Mathy, Olivier Arnaiz, Julita Gruchota, Robert Gromadka, Mireille Bétermier, Sandra Duharcourt, Cyril Denby Wilkes, Jacek K NowakAbstract:Because of their Nuclear Dimorphism, ciliates provide a unique opportunity to study the role of non-coding RNAs (ncRNAs) in the communication between germline and somatic lineages. In these unicellular eukaryotes, a new somatic nucleus develops at each sexual cycle from a copy of the zygotic (germline) nucleus, while the old somatic nucleus degenerates. In the ciliate Paramecium tetraurelia, the genome is massively rearranged during this process through the reproducible elimination of repeated sequences and the precise excision of over 45,000 short, single-copy Internal Eliminated Sequences (IESs). Different types of ncRNAs resulting from genome-wide transcription were shown to be involved in the epigenetic regulation of genome rearrangements. To understand how ncRNAs are produced from the entire genome, we have focused on a homolog of the TFIIS elongation factor, which regulates RNA polymerase II transcriptional pausing. Six TFIIS-paralogs, representing four distinct families, can be found in P. tetraurelia genome. Using RNA interference, we showed that TFIIS4, which encodes a development-specific TFIIS protein, is essential for the formation of a functional somatic genome. Molecular analyses and high-throughput DNA sequencing upon TFIIS4 RNAi demonstrated that TFIIS4 is involved in all kinds of genome rearrangements, including excision of ~48% of IESs. Localization of a GFP-TFIIS4 fusion revealed that TFIIS4 appears specifically in the new somatic nucleus at an early developmental stage, before IES excision. RT-PCR experiments showed that TFIIS4 is necessary for the synthesis of IES-containing non-coding transcripts. We propose that these IES+ transcripts originate from the developing somatic nucleus and serve as pairing substrates for germline-specific short RNAs that target elimination of their homologous sequences. Our study, therefore, connects the onset of zygotic non coding transcription to the control of genome plasticity in Paramecium, and establishes for the first time a specific role of TFIIS in non-coding transcription in eukaryotes.
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The Paramecium Germline Genome Provides a Niche for Intragenic Parasitic DNA: Evolutionary Dynamics of Internal Eliminated Sequences
PLOS Genetics, 2012Co-Authors: Olivier Arnaiz, Nathalie Mathy, Céline Baudry, Sophie Malinsky, Cyril Denby Wilkes, Benjamin E. Lauderdale, Jean-marc Aury, Olivier Garnier, Karine Labadie, Anne Le MouëlAbstract:Insertions of parasitic DNA within coding sequences are usually deleterious and are generally counter-selected during evolution. Thanks to Nuclear Dimorphism, ciliates provide unique models to study the fate of such insertions. Their germline genome undergoes extensive rearrangements during development of a new somatic macronucleus from the germline micronucleus following sexual events. In Paramecium, these rearrangements include precise excision of unique-copy Internal Eliminated Sequences (IES) from the somatic DNA, requiring the activity of a domesticated piggyBac transposase, PiggyMac. We have sequenced Paramecium tetraurelia germline DNA, establishing a genome-wide catalogue of ∼45,000 IESs, in order to gain insight into their evolutionary origin and excision mechanism. We obtained direct evidence that PiggyMac is required for excision of all IESs. Homology with known P. tetraurelia Tc1/mariner transposons, described here, indicates that at least a fraction of IESs derive from these elements. Most IES insertions occurred before a recent whole-genome duplication that preceded diversification of the P. aurelia species complex, but IES invasion of the Paramecium genome appears to be an ongoing process. Once inserted, IESs decay rapidly by accumulation of deletions and point substitutions. Over 90% of the IESs are shorter than 150 bp and present a remarkable size distribution with a ∼10 bp periodicity, corresponding to the helical repeat of double-stranded DNA and suggesting DNA loop formation during assembly of a transpososome-like excision complex. IESs are equally frequent within and between coding sequences; however, excision is not 100% efficient and there is selective pressure against IES insertions, in particular within highly expressed genes. We discuss the possibility that ancient domestication of a piggyBac transposase favored subsequent propagation of transposons throughout the germline by allowing insertions in coding sequences, a fraction of the genome in which parasitic DNA is not usually tolerated.
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transposon invasion of the paramecium germline genome countered by a domesticated piggybac transposase and the nhej pathway
International Journal of Evolutionary Biology, 2012Co-Authors: Nathalie Mathy, Emeline Dubois, Julien Bischerour, Antoine Marmignon, Vinciane Regnier, Mireille BétermierAbstract:Sequences related to transposons constitute a large fraction of extant genomes, but insertions within coding sequences have generally not been tolerated during evolution. Thanks to their unique Nuclear Dimorphism and to their original mechanism of programmed DNA elimination from their somatic nucleus (macronucleus), ciliates are emerging model organisms for the study of the impact of transposable elements on genomes. The germline genome of the ciliate Paramecium, located in its micronucleus, contains thousands of short intervening sequences, the IESs, which interrupt 47% of genes. Recent data provided support to the hypothesis that an evolutionary link exists between Paramecium IESs and Tc1/mariner transposons. During development of the macronucleus, IESs are excised precisely thanks to the coordinated action of PiggyMac, a domesticated piggyBac transposase, and of the NHEJ double-strand break repair pathway. A PiggyMac homolog is also required for developmentally programmed DNA elimination in another ciliate, Tetrahymena. Here, we present an overview of the life cycle of these unicellular eukaryotes and of the developmentally programmed genome rearrangements that take place at each sexual cycle. We discuss how ancient domestication of a piggyBac transposase might have allowed Tc1/mariner elements to spread throughout the germline genome of Paramecium, without strong counterselection against insertion within genes.
