137 results on '"Dirks, Ron"'
Search Results
102. The Sequence of Regulatory Events Controlling the Expression of the γD-crystallin Gene during Fibroblast Growth Factor-Mediated Rat Lens Fiber Cell Differentiation
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Dirks, Ron P.H., primary, Klok, Erik Jan, additional, van Genesen, Siebe T., additional, Schoenmakers, John G.G., additional, and Lubsen, Nicolette H., additional
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- 1996
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103. A novel humanc-sismRNA species is transcribed from a promoter inc-sisintron 1 and contains the code for an alternative PDGF B-like protein
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Dirks, Ron P.H., primary, Onnekink, Carla, additional, Jansen, Hans J., additional, de Jong, Aard, additional, and Bloemers, Henri P.J., additional
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- 1995
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104. In vivofootprinting and functional analysis of the human c-sis/PDGF B gene promoter provides evidence for two binding sites for transcriptional activators
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Dirks, Ron P. H., primary, Jansen, Hans J., additional, van Gerven, Bart, additional, Onnekink, Carla, additional, and Bloemers, Henri P. J., additional
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- 1995
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105. Testing Tuberculosis Drug Efficacy in a Zebrafish High-Throughput Translational Medicine Screen
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Ordas, Anita, Raterink, Robert-Jan, Cunningham, Fraser, Jansen, Hans J., Wiweger, Malgorzata I., Jong-Raadsen, Susanne, Bos, Sabine, Bates, Robert H., Barros, David, Meijer, Annemarie H., Vreeken, Rob J., Ballell-Pages, Lluís, Dirks, Ron P., Hankemeier, Thomas, and Spaink, Herman P.
- Abstract
ABSTRACTThe translational value of zebrafish high-throughput screens can be improved when more knowledge is available on uptake characteristics of potential drugs. We investigated reference antibiotics and 15 preclinical compounds in a translational zebrafish-rodent screening system for tuberculosis. As a major advance, we have developed a new tool for testing drug uptake in the zebrafish model. This is important, because despite the many applications of assessing drug efficacy in zebrafish research, the current methods for measuring uptake using mass spectrometry do not take into account the possible adherence of drugs to the larval surface. Our approach combines nanoliter sampling from the yolk using a microneedle, followed by mass spectrometric analysis. To date, no single physicochemical property has been identified to accurately predict compound uptake; our method offers a great possibility to monitor how any novel compound behaves within the system. We have correlated the uptake data with high-throughput drug-screening data from Mycobacterium marinum-infected zebrafish larvae. As a result, we present an improved zebrafish larva drug-screening platform which offers new insights into drug efficacy and identifies potential false negatives and drugs that are effective in zebrafish and rodents. We demonstrate that this improved zebrafish drug-screening platform can complement conventional models of in vivoMycobacterium tuberculosis-infected rodent assays. The detailed comparison of two vertebrate systems, fish and rodent, may give more predictive value for efficacy of drugs in humans.
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- 2014
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106. DNase-I-hypersensitive sites located far upstream of the human c-sis/PDGF-B gene comap with transcriptional enhancers and a silencer and are preceded by (part of) a new transcription unit
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DIRKS, Ron P. H., primary, JANSEN, Hans J., additional, ONNEKINK, Carla, additional, JONGE, Rob J. A., additional, and BLOEMERS, Henri P. J., additional
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- 1993
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107. Localization and functional analysis of DNase-I-hypersensitive sites in the human c-sis/PDGF-B gene transcription unit and its flanking regions
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DIRKS, Ron P. H., primary, JANSEN, Hans J., additional, GERRITSMA, Jort, additional, ONNEKINK, Carla, additional, and BLOEMERS, Henri P. J., additional
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- 1993
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108. Splicing of the platelet-derived-growth-factor A-chain mRNA in human malignant mesothelioma cell lines and regulation of its expression
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LANGERAK, Anthonie W., primary, DIRKS, Ron P. H., additional, and VERSNEL, Marjan A., additional
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- 1992
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109. Cell-Autonomous TrkB Signaling in Presynaptic Retinal Ganglion Cells Mediates Axon Arbor Growth and Synapse Maturation during the Establishment of Retinotectal Synaptic Connectivity. .
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Marshak, Sonya, Nikolakopoulou, Angeliki Maria, Dirks, Ron, Martens, Gerard J., and Cohen-Cory, Susana
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RETINAL ganglion cells ,AXONS ,SYNAPSES ,XENOPUS laevis ,GREEN fluorescent protein - Abstract
BDNF contributes to the activity-dependent establishment and refinement of visual connectivity. In Xenopus, BDNF applications in the optic tectum influence retinal ganglion cell (RGC) axon branching and promote synapse formation and stabilization. The expression patterns of BDNF and TrkB suggest that BDNF specifically regulates the maturation of RGC axons at the target. It is possible, however, that BDNF modulates retinotectal synaptic connectivity by differentially influencing presynaptic RGC axons and postsynaptic tectal cells. Here, we combined single-cell expression of a dominant-negative TrkB-enhanced green fluorescent protein (GFP) fusion protein with confocal microscopy imaging in live Xenopus tadpoles to differentiate between presynaptic and postsynaptic actions of BDNF. Disruption of TrkB signaling in individual RGCs influenced the branching and synaptic maturation of presynaptic axon arbors. Specifically, GFP-TrkB.T1 overexpression increased the proportion of axons with immature, growth cone-like morphology, decreased axon branch stability, and increased axon arbor degeneration. In addition, GFP-TrkB.T1 overexpression reduced the number of red fluorescent protein-synaptobrevin-labeled presynaptic specializations per axon terminal. In contrast, overexpression of GFP-TrkB.T1 in tectal neurons did not alter synaptic number or the morphology or dynamic behavior of their dendritic arbors. Electron microscopy analysis revealed a significant decrease in the number of mature synaptic profiles and in the number of docked synaptic vesicles at retinotectal synapses made by RGC axons expressing GFP-TrkB.T1. Together, our results demonstrate that presynaptic TrkB signaling in RGCs is a key determinant in the establishment of visual connectivity and indicate that changes in tectal neuron synaptic connectivity are secondary to the BDNF-elicited enhanced stability and growth of presynaptic RGCs. [ABSTRACT FROM AUTHOR]
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- 2007
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110. Sequence and Functional Conservation of the Intergenic Region Between the Head-to-Head Genes Encoding the Small Heat Shock Proteins aB-Crystallin and HspB2 in the Mammalian Lineage.
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Doerwald, Linda, van Rheede, Teun, Dirks, Ron P., Madsen, Ole, Rexwinkel, Remco, van Genesen, Siebe T., Martens, Gerard J., de Jong, Wilfried W., and Lu, Nicolette H.
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HEAT shock proteins ,PROTEINS ,GENETIC code ,NUCLEOTIDE sequence ,MOLECULAR evolution ,ORIGIN of life ,EVOLUTIONARY theories ,MOLECULAR biology - Abstract
An unexpected feature of the large mammalian genome is the frequent occurrence of closely linked head-to-head gene pairs. Close apposition of such gene pairs has been suggested to be due to sharing of regulatory elements. We show here that the head-to-head gene pair encoding two small heat shock proteins, aB-crystallin and HspB2, is closely linked in all major mammalian clades, suggesting that this close linkage is of selective advantage. Yet aB-crystallin is abundantly expressed in lens and muscle and in response to a heat shock, while HspB2 is abundant only in muscle and not upregulated by a heat shock. The intergenic distance between the genes for these two proteins in mammals ranges from 645 bp (platypus) to 1069 bp (opossum), with an average of about 900 bp; in chicken the distance was the same as in duck (1.6 kb). Phylogenetic footprinting and sequence alignment identified a number of conserved sequence elements close to the HspB2 promoter and two farther upstream. All known regulatory elements of the mouse aB-crystallin promoter are conserved, except in platypus and birds. The lens-specific region 1 (LSR1) and the heat shock elements (HSEs) lack in birds; in platypus the LSR1 is reduced to a Pax-6 site, while the Pax-6 site in LSR2 and a HSE are absent. Most likely the primordial mammalian aB-crystallin promoter had two LSRs and two HSEs. In transfection experiments the platypus aB-crystallin promoter retained heat shock responsiveness and lens expression. It also directed lens expression inXenopus laevistransgenes, as did the HspB2 promoter of rat or blind mole rat. Deletion of the middle of the intergenic region including the upstream enhancer affected the activity of both the rat aB-crystallin and the HspB2 promoters, suggesting sharing of the enhancer region by the two promoters. [ABSTRACT FROM AUTHOR]
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- 2004
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111. The cooperation between two silencers creates an enhancer element that controls both the lens-preferred and the differentiantion stage-specific expression of the rat βB2;-crystallin gene.
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Dirks, Ron P. H., Kraft, Harry J., van Genesen, Siebe T., Klok, Erik J., Pfundt, Rolph, Schoenmakers, John G. G., and Lubsen, Nicolette H.
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CELLS , *GROWTH factors , *CYTOKINES , *CELLULAR immunity , *CHEMOKINES , *PEPTIDES - Abstract
The rat βB2-crystallin gene is active only during a specific stage of the differentiation of rat lens fibre cells directed by basic fibroblast growth factor. The regulatory elements that determine the transient activity of this gene are located in the -750/-123 region and in the first intron. Singly, these elements act as silencers, together they constitute an enhancer that is active only during the specific differentiation stage. An additional silencer is found between -123 and -77. The proximal promoter region contains a Pax-6 binding site at -65/-51 . In vitro, binding to this site could be detected but, according to in vivo footprinting experiments, this site is not occupied in the endogenous gene. Furthermore, co-expression of Pax-6 did not enhance promoter activity. Finally, mutation or deletion of this site did not affect promoter activity; the region -37/+10 sufficed for basal promoter activity. The cooperation between the -750/-123 region and the first intron of the βB2-crystanin gene not only determines the differentiation stage-specific activity of the gene, but also contributes to the highly increased expression in lens cells compared with non-lens cells. [ABSTRACT FROM AUTHOR]
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- 1996
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112. DNase-1-hypersensitive sites located far upstream of the human c-sis/PDGF-B gene comap with transcriptional enhancers and a silencer are preceded by (part of) a new transcription unit.
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Dirks, Ron P. H., Jansen, Hans J., Onnekink, Carla, de Jonge, Rob J. A., and Bloemers, Henri P. J.
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GENETIC code , *BLOOD platelets , *GROWTH factors , *BONE marrow , *MYELOID leukemia , *GENETIC transcription , *MESSENGER RNA , *PHORBOL esters , *PROMOTERS (Genetics) - Abstract
The human c-sis gene encodes the B chain of platelet-derived growth factor (PDGF), a potent mitogen for cultured cells of mesenchymal origin. PDGF is stored in the α-granules of blood platelets, which are derived from bone marrow megakaryocytes and lack transcriptional machinery. Human myeloid leukemia cell line K562 can be used as a model for megakaryocytes. Phorbol-estermediated megakaryocytic ddifferentiation of K562 cells is accompanied by more than 200-fold increase in the c-sis mRNA level. We have now localized transcriptional enhancers at −8.6 kb and −9.9 kb relative to the human c-sis gene transcription start site. The enhancer at −8.6 kb increases activity of the c0is promoter by 40–60-fold specifically in K562 cells and comaps with a Dnase-I-hypersensitivity (DH) site. The enhancer at − 9.9 kb increases c-sis promoter activity by 5–10-fold in K562 cells and DH at that site accompanies phorbol-ester-induced megakaryocytic differentiation. In phorbol-ester-treated K562 cells the two enhancers may be negatively influenced by a silencer that comaps with DH at −10.7/−11.0 kb. Reporter gene analysis predicted that combined activity of the upstream enhancers and the c-sis promoter may result in 100–1000-fold higher promoter activity in phorbol-ester-treated K562 cells compared with untreated cells, which can fully explain the more than 200-fold increase in c-sis mRNA level. DH at −8.6 kb and −9.9 kb was also detected in human fibroblasts and in the carcinoma cell lines HeLa and PC3, which express, respectively, undetectable, low and high levels of c-sis mRNA. Although the individual DH sites displayed 4—10-fold enhancer activity in all these cells, they lost most of their biological activity when combined in a larger fragment. In addition we localized (part of) a new transcription unit at approximately 13 kb upstream of the c-sis transcription start site. The corresponding 0.45-kb sis upstream region (sur) transcript is constitutively expressed in all cell lines examined. The expression of the sur transcript is independent of the expression of c-sis mRNA and of the pattern of DH sites far upstream of the c-sis gene. Thus, at present, there is no indication that the upstream DH sites are involved in regulation of expression of the sur gene. [ABSTRACT FROM AUTHOR]
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- 1993
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113. A novel human c-sis mRNA species is transcribed from a promoter in c-sis intron 1 and contains the code for an alternative PDGF B-like protein.
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Dirks, Ron P.H., Onnekink, Carla, Jansen, Hans J., de Jong, Aard, and Bloemers, Henri P.J.
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- 1995
114. In vivo footprinting and functional analysis of the human c-sis/PDGF B gene promoter provides evidence for two binding sites for transcriptional activators.
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Dirks, Ron P. H., Jansen, Hans J., van Gerven, Bart, Onnekink, Carla, and Bloemers, Henri P. J.
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- 1995
115. Supplementary text S1-S5; Tables S1-S2; Figures S1-S16 from A 180 Myr-old female-specific genome region in sturgeon reveals the oldest known vertebrate sex determining system with undifferentiated sex chromosomes
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Kuhl, Heiner, Guiguen, Yann, Höhne, Christin, Kreuz, Eva, Du, Kang, Klopp, Christophe, Lopez-Roques, Céline, Yebra-Pimentel, Elena Santidrian, Ciorpac, Mitica, Gessner, Jörn, Holostenco, Daniela, Kleiner, Wibke, Kohlmann, Klaus, Lamatsch, Dunja K., Prokopov, Dmitry, Bestin, Anastasia, Bonpunt, Emmanuel, Debeuf, Bastien, Haffray, Pierrick, Morvezen, Romain, Patrice, Pierre, Suciu, Radu, Dirks, Ron, Wuertz, Sven, Kloas, Werner, Schartl, Manfred, and Stöck, Matthias
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14. Life underwater - Abstract
Several hypotheses explain the prevalence of undifferentiated sex chromosomes in poikilothermic vertebrates. Turnovers change the master sex determination gene, the sex chromosome or the sex determination system (e.g. XY to WZ). Jumping master genes stay main triggers but translocate to other chromosomes. Occasional recombination (e.g. in sex-reversed females) prevents sex chromosome degeneration. Recent research has uncovered conserved heteromorphic or even homomorphic sex chromosomes in several clades of non-avian and non-mammalian vertebrates. Sex determination in sturgeons (Acipenseridae) has been a long-standing basic biological question, linked to economical demands by the caviar-producing aquaculture. Here, we report the discovery of a sex-specific sequence from sterlet (Acipenser ruthenus). Using chromosome-scale assemblies and pool-sequencing, we first identified an approximately 16 kb female-specific region. We developed a PCR-genotyping test, yielding female-specific products in six species, spanning the entire phylogeny with the most divergent extant lineages (A. sturio, A. oxyrinchus versus A. ruthenus, Huso huso), stemming from an ancient tetraploidization. Similar results were obtained in two octoploid species (A. gueldenstaedtii, A. baerii). Conservation of a female-specific sequence for a long period, representing 180 Myr of sturgeon evolution, and across at least one polyploidization event, raises many interesting biological questions. We discuss a conserved undifferentiated sex chromosome system with a ZZ/ZW-mode of sex determination and potential alternatives.This article is part of the theme issue ‘Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part I)’.
116. Supplementary text S1-S5; Tables S1-S2; Figures S1-S16 from A 180 Myr-old female-specific genome region in sturgeon reveals the oldest known vertebrate sex determining system with undifferentiated sex chromosomes
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Kuhl, Heiner, Guiguen, Yann, Höhne, Christin, Kreuz, Eva, Du, Kang, Klopp, Christophe, Lopez-Roques, Céline, Yebra-Pimentel, Elena Santidrian, Ciorpac, Mitica, Gessner, Jörn, Holostenco, Daniela, Kleiner, Wibke, Kohlmann, Klaus, Lamatsch, Dunja K., Prokopov, Dmitry, Bestin, Anastasia, Bonpunt, Emmanuel, Debeuf, Bastien, Haffray, Pierrick, Morvezen, Romain, Patrice, Pierre, Suciu, Radu, Dirks, Ron, Wuertz, Sven, Kloas, Werner, Schartl, Manfred, and Stöck, Matthias
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14. Life underwater - Abstract
Several hypotheses explain the prevalence of undifferentiated sex chromosomes in poikilothermic vertebrates. Turnovers change the master sex determination gene, the sex chromosome or the sex determination system (e.g. XY to WZ). Jumping master genes stay main triggers but translocate to other chromosomes. Occasional recombination (e.g. in sex-reversed females) prevents sex chromosome degeneration. Recent research has uncovered conserved heteromorphic or even homomorphic sex chromosomes in several clades of non-avian and non-mammalian vertebrates. Sex determination in sturgeons (Acipenseridae) has been a long-standing basic biological question, linked to economical demands by the caviar-producing aquaculture. Here, we report the discovery of a sex-specific sequence from sterlet (Acipenser ruthenus). Using chromosome-scale assemblies and pool-sequencing, we first identified an approximately 16 kb female-specific region. We developed a PCR-genotyping test, yielding female-specific products in six species, spanning the entire phylogeny with the most divergent extant lineages (A. sturio, A. oxyrinchus versus A. ruthenus, Huso huso), stemming from an ancient tetraploidization. Similar results were obtained in two octoploid species (A. gueldenstaedtii, A. baerii). Conservation of a female-specific sequence for a long period, representing 180 Myr of sturgeon evolution, and across at least one polyploidization event, raises many interesting biological questions. We discuss a conserved undifferentiated sex chromosome system with a ZZ/ZW-mode of sex determination and potential alternatives.This article is part of the theme issue ‘Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part I)’.
117. Supplementary text S1-S5; Tables S1-S2; Figures S1-S16 from A 180 Myr-old female-specific genome region in sturgeon reveals the oldest known vertebrate sex determining system with undifferentiated sex chromosomes
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Kuhl, Heiner, Guiguen, Yann, Höhne, Christin, Kreuz, Eva, Du, Kang, Klopp, Christophe, Lopez-Roques, Céline, Yebra-Pimentel, Elena Santidrian, Ciorpac, Mitica, Gessner, Jörn, Holostenco, Daniela, Kleiner, Wibke, Kohlmann, Klaus, Lamatsch, Dunja K., Prokopov, Dmitry, Bestin, Anastasia, Bonpunt, Emmanuel, Debeuf, Bastien, Haffray, Pierrick, Morvezen, Romain, Patrice, Pierre, Suciu, Radu, Dirks, Ron, Wuertz, Sven, Kloas, Werner, Schartl, Manfred, and Stöck, Matthias
- Subjects
14. Life underwater - Abstract
Several hypotheses explain the prevalence of undifferentiated sex chromosomes in poikilothermic vertebrates. Turnovers change the master sex determination gene, the sex chromosome or the sex determination system (e.g. XY to WZ). Jumping master genes stay main triggers but translocate to other chromosomes. Occasional recombination (e.g. in sex-reversed females) prevents sex chromosome degeneration. Recent research has uncovered conserved heteromorphic or even homomorphic sex chromosomes in several clades of non-avian and non-mammalian vertebrates. Sex determination in sturgeons (Acipenseridae) has been a long-standing basic biological question, linked to economical demands by the caviar-producing aquaculture. Here, we report the discovery of a sex-specific sequence from sterlet (Acipenser ruthenus). Using chromosome-scale assemblies and pool-sequencing, we first identified an approximately 16 kb female-specific region. We developed a PCR-genotyping test, yielding female-specific products in six species, spanning the entire phylogeny with the most divergent extant lineages (A. sturio, A. oxyrinchus versus A. ruthenus, Huso huso), stemming from an ancient tetraploidization. Similar results were obtained in two octoploid species (A. gueldenstaedtii, A. baerii). Conservation of a female-specific sequence for a long period, representing 180 Myr of sturgeon evolution, and across at least one polyploidization event, raises many interesting biological questions. We discuss a conserved undifferentiated sex chromosome system with a ZZ/ZW-mode of sex determination and potential alternatives.This article is part of the theme issue ‘Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part I)’.
118. Nucleotide sequence and expression of a β-tubulin gene from Plasmodium falciparum, a malarial parasite of man
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Wesseling, John G., primary, Dirks, Ron, additional, Smits, Man A., additional, and Schoenmakers, John G.G., additional
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- 1989
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119. New genomics and transcriptomics tools toward improving conservation strategies for sturgeons
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Santidrian Yebra-Pimentel, Elena Maria, Dirks, Ron P., Weltzien, Finn-Arne, and Dufour, Sylvie
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HSP ,Acipenser - Abstract
Sturgeons (Family Acipenseridae) are one of the largest and most primitive fish families on Earth. Although they have always been typically distributed throughout the Northern Hemisphere, during the last decades wild populations have declined due to anthropogenic factors such as overfishing, poaching, pollution, and habitat loss. The situation is particularly dramatic for Atlantic sturgeon, one of the most ancient species among the family, which is currently extinct in Europe. In order to reintroduce the Atlantic sturgeon in Europe, several Baltic countries have been working together for more than two decades to build an ex-situ broodstock locally with fish derived from Canada, where the populations are not threatened, and releasing juveniles into the rivers flowing to the Baltic Sea.However, rearing fish aimed for restocking in the same manner as aquacultured fish has been shown to impact the post-release survival of juveniles in the long term. For example, in aquaculture conditions, fish are often maintained in high densities, at constant photoperiod and temperature conditions, and feeding on commercial pellets, leaving them cognitively naïve when released into natural environments. Additionally, increased water temperatures due to global warming have a strong influence on the geographic distribution of the species, resulting in local extinctions and population shifts. Although the effect of heat and cold stress on the juvenile and embryonic development have been assessed for several sturgeon species, most studies on gene expression have looked at a very limited number of genes due to the lack of sequence information and genomic resources. Also, most studies use other sturgeon species such as Siberian, Japanese, and white sturgeons, which are widespread aquacultured species. Exposing fish to temperatures higher than the optimal can trigger phenotypic adaptations leading to increase thermotolerance and potentially improve postrelease survival, however the impact of temperature-training protocols on the response to a subsequent heat shock has not yet been assessed in sturgeons. Therefore, the main aim of this thesis was to generate genomic and transcriptomic resources for Atlantic sturgeon, which are essential to improve and promote research in many fields, such as ecology, physiology and evolutionary studies. Moreover, it provides a reference for RNAseq-mediated transcriptome mapping. Additionally, we have used these resources to develop and evaluate the impact of novel rearing techniques toward improving restoration success, focusing on temperature training. First, we have assembled a high-quality de novo transcriptome, made an inventory of all the heat shock protein (HSP) gene family members and exposed a cell line derived from Atlantic sturgeon larvae to a moderate and severe heat shock in order to identify all heat-responsive genes using an RNAseq approach (Paper I). We found 76 HSP genes in the Atlantic sturgeon transcriptome, only 16 of which were responsive to at least one of the applied heat shock protocols, and only 5 of which were consistently upregulated after both moderate and severe heat shock at all the tested timepoints. After building the reference transcriptome and annotating all the HSP genes, we have evaluated the differences in liver transcriptome between temperature-trained and nontrained juveniles upon exposure to a new heat shock (Paper II). After four weeks of treatment, fish exposed to temperature training showed between 2 to 4 fold less dysregulated genes in response to a new heat shock than the non-trained group, indicating their improved ability to maintain transcriptomic homeostasis during a new heat shock. Again, like in the in vitro experiment, very few of the annotated HSP genes were dysregulated in response to heat shock in the liver transcriptome, namely hspa1, hspc1 and dnajb4. Overall, the response to heat shock in the liver transcriptome was milder than the in vitro response, which is likely a consequence of the activation of compensatory mechanisms. These mechanisms include the neuroendocrine system and result in increased tissue protection and thermogenic capacities, especially in the trained fish. We therefore propose that temperature-training protocols like the one tested in this thesis should be included in the set of new rearing techniques for fish used for restocking; however, other protocols should be investigated. Since the main bottleneck in the evaluation of the effect of such training is the lack of sequence information and a reference genome for RNAseq experiments, we have additionally assembled a reference genome for Atlantic sturgeon using a combination of short and long-read sequencing technologies (Paper III). The assembled genome provides for the first time clear evidence of a sturgeon-specific whole-genome duplication event (SR), independent from the American paddlefish (Polyodon spathula), which is the main representative of the sister Family (Polyodontidae) within the same Order (Acipensiformes). The presence of duplicated Hox clusters, together with synteny and phylogenetic studies of these developmental genes, and the results of microsatellite loci analysis, suggests that sturgeons have a paleotetraploid origin, and that a rediploidization process is still ongoing. In summary, the results presented in this thesis advance the field of sturgeon research. We hypothesized that temperature training has a positive effect during the exposure to a subsequent heat shock, but its potential to improve post-release survival in the long term should still be assessed. We therefore suggest that future work should be aimed at the optimization of rearing methods for stocking programs and that a reference genome should be used. Stør (Familie Acipenseridae) er blant de største og mest primitive familier av benfisk. Mens de opprinnelig var utbredt over hele den nordlige halvkule, er mange populasjoner nå kritisk truet på grunn av menneskeskapte faktorer som overfiske, forurensning og tap av habitat. Situasjonen er spesielt dramatisk for atlantisk stør, en av de eldste artene i familien, som er utdødd i Europa. For å gjeninnføre atlantisk stør, har flere baltiske land arbeidet for å etablere en ex situ stamfiskpopulasjon basert på fisk fra Canada (hvor bestanden ikke er truet), for produksjon av yngel til utsetting i baltiske vassdrag. Imidlertid gir produksjon av settefisk etter samme prinsipper som for oppdrettsfisk svært lav overlevelse i naturen. Eksempelvis vil høy tetthet, konstant fotoperiode og vanntemperatur, og fôring med pellets til faste tider gi en kognitivt naiv fisk som ikke klarer seg i det fri. I tillegg kommer økte vanntemperaturer som resultat av global oppvarming, og som har stor innvirkning på den geografiske fordelingen av arter, som igjen resulterer i lokal utryddelse og forflytning av populasjoner. Selv om effekter av vanntemperatur har blitt undersøkt på embryonal- og yngelutviklingen i flere størarter, har de fleste studier fokusert på et lite sett av gener fordi genomiske ressurser har manglet. I tillegg er de fleste studiene utført på andre arter som er vanlig i akvakultur, slik som sibirsk, japansk og hvit stør. Eksponering til vanntemperaturer som er høyere enn artens optimum, kan utløse fenotypiske tilpasninger som fører til økt termotoleranse og potensielt forbedre overlevelse i naturen. Men effekten av slike temperaturtreningsprotokoller på responsen på et påfølgende varmesjokk er ikke studert i stør. Hovedmålet med denne avhandlingen var å generere genomiske og transkriptomiske ressurser for atlantisk stør, som en viktig faktor for å forbedre og fremme forskning innen økologi, fysiologi og evolusjon. Videre gir avhandlingen en referanse for RNAseq-mediert transkriptomkartlegging. Disse ressursene er så benyttet til å utvikle og evaluere virkningen av nye oppdrettsteknikker for settefisk, med fokus på temperaturtrening. Først har vi satt sammen et høykvalitets de novo transkriptom, deretter karakterisert genfamilien av varmesjokkproteiner (HSP), og så eksponert en cellelinje avledet fra atlantiske størlarver for et moderat og et kraftig varmesjokk for å identifisere alle varmeresponsive gener ved bruk av RNAseq (Artikkel I). Vi fant 76 HSP-gener i transkriptomene fra atlantisk stør, hvorav 16 responderte på minst en av de testede varmesjokkprotokollene, og bare 5 av disse ble konsekvent oppregulert etter både moderat og kraftig varmesjokk ved alle testede tidspunkt. Etter å ha bygget referansetranskriptomet og karakterisert alle HSP-gener, evaluerte vi forskjellene i levertranskriptom mellom temperaturtrent og ikke-trent yngel ved eksponering for et nytt varmesjokk (Artikkel II). Etter fire ukers behandling viste fisk som ble utsatt for temperaturtrening mellom 2-4 ganger færre dysregulerte gener som svar på et nytt varmesjokk sammenlignet med den ikke-trente gruppen. Dette antyder en forbedret evne til å opprettholde transkriptomisk homeostase under et nytt varmesjokk. I likhet med in vitro eksperimentet var svært få HSP-gener dysregulert som respons på varmesjokk i levertranskriptomet, nemlig hspa1, hspc1 og dnajb4. Totalt sett var responsen på varmesjokk i levertranskriptomet mildere enn in vitro responsen, noe som sannsynligvis skyldes ulike kompensatoriske mekanismer. Disse inkluderer det nevroendokrine systemet og resulterer i økt vevsbeskyttelse og termogen kapasitet, spesielt i trent fisk. Selv om andre protokoller bør undersøkes nærmere, foreslår vi at protokoller for temperaturtrening lik den som ble testet i denne avhandlingen bør vurderes i nye oppdrettsprotokoller for settefiskproduksjon av stør. Siden den viktigste flaskehalsen i evalueringen av effekten av slik trening er mangelen på sekvensinformasjon og et referansegenom for RNAseq-eksperimenter, har vi i tillegg produsert et referansegenom for atlantisk stør ved bruk av ulike sekvenseringsteknologier for korte og lange reads (Artikkel III). Genomet gir for første gang klare bevis for en størspesifikk helgenomdupliseringshendelse (SR), uavhengig av spadestør (Polyodon spathula), som er hovedrepresentanten for søsterfamilien (Polyodontidae) innenfor samme Orden (Acipensiformes). Tilstedeværelsen av dupliserte klynger av hox-gener som er sentrale i tidlig utvikling, i tillegg til hox-gen synteni og fylogeni, og mikrosatellitt loci-analyser, antyder at stør har en paleotetraploid opprinnelse, og at en rediploidiseringsprosess fortsatt pågår. Oppsummert vil de forbedrete genomiske og transkriptomiske verktøy presentert i denne avhandlingen åpne for nye muligheter innen størforskning. Videre har temperaturtrening en positiv effekt ved eksponeringen til varmesjokk, men potensialet for temperaturtrening og dermed økt overlevelse ved utsett bør undersøkes videre. Os esturións (Familia Acipenseridae) pertencen a unha das familias de peixes mais grandes e primitivas da Terra. Aínda que sempre estiveron distribuídos no hemisferio Norte, nas últimas décadas as poboacións salvaxes teñen diminuido debido a factores antropoxénicos como a sobrepesca, a caza furtiva, a contaminación e a perda do hábitat. A situación é especialmente dramática para o esturión Atlántico, unha das especies máis antigas da familia que se atopa extinta en Europa na actualidade. Co fin de reintroducir o esturión Atlántico en Europa, varios países bálticos levan traballando xuntos durante máis de dúas décadas para construír un núcleo reprodutor con peixes derivados de Canadá, onde a poboación non esta ameazada, e liberar xuvenís nos ríos que flúen ao Mar Báltico. Non obstante, cultivar peixe destinado ao repoboamento utilizando as mesmas técnicas típicamente utilizadas en acuicultura afecta negativamente á supervivencia a longo prazo. Por exemplo, en condicións de acuicultura os peixes adoitan producirse en altas densidades, baixo condicións abióticas constantes (fotoperíodo e temperatura), e aliméntanse de pellets comerciais, producindo animais congitivamente inxenuos cuando son libertados no ambiente natural. Ademais, o aumento das temperaturas da auga debido ao quentamento global ten unha forte influencia na distribución xeográfica das especies, dando lugar a extincións locais. Aínda que os efectos do estrés térmico no desenvolvemento embrionario e etapas xuvenís xa teñen sido avaliados en varias especies de esturións, a maioría dos estudos de expresión xénica céntranse en un número moi limitado de xenes debido á falta de recursos xenómicos. Ademáis, a maioría dos estudos utilizan outras especies de esturións como o branco, siberiano ou xaponés, mais comúns en acuicultura. A exposición dos peixes a temperaturas superiores ás óptimas pode desecandear adaptacións fenotípicas resultando nun incremento da tolerancia térmica e potencialmente unha mellora da supervivencia no hábitat onde son libertados. Sen embargo, os efectos do réxime de temperatura utilizado durante a cría na resposta a un choque de calor posterior non teñen sido avaliados. Por todo isto, o obxectivo principal desta tese foi xerar recursos xenómicos e transcriptómicos para o esturión Atlántico, esenciais para mellorar e promover a investigación en moitos campos da ciencia como a ecoloxía, fisioloxía e a xenómica evolutiva, ademais de fornecer unha referencia para o mapeado do transcriptoma. Ademais, empregamos estes recursos para desenvolver e avaliar o impacto de novas técnicas de cultivo para mellorar o proceso de repoboación, centrándose na xeración de individuos termotolerantes. En primeiro lugar, temos ensamblado un transcriptoma de alta calidade, fixemos un inventario de todos os membros da familia das proteínas de choque térmico (HSP) e expuxemos unha liña celular isolada a partir de larvas disgregadas de esturión Atlántico a un choque de calor moderado e severo para identificar xenes sensibles ó calor (Artigo I). Atopamos 76 HSP no transcriptoma de esturión Atlántico, dos cuáis só 16 foron sensibles a polo menos un dos protocolos de choque de calor avalidado, e 5 foron sensibles a ambos choques térmicos idenpendentemene do momento da amostraxe. Utilizando as secuencias dos xens HSP e o trancriptoma de esturión ensamblado no Artigo I como referencia, temos avaliado as diferenzas no transcriptoma hepático en resposta a un choque térmico entre xuvenís criados baixo un réxime de temperatura constante e en réxime fluctuante (Artigo II). Despois de catro semanas de tratamento, os peixes criados en réxime de temperatura fluctuante mostraron entre 2 e 4 veces menos xens diferencialmente expresados en resposta a un novo choque térmico que os peixes criados en réxime de temperatura constante, indicando a súa mellor capacidade para manter a homeostase transcriptómica durante un novo choque térmico. Como xa indicaron os resultados in vitro, moi poucos HSP foron diferencialmente expresados en resposta ó choque de calor no transcriptoma hepático, concretamente hspa1, hspc1 e dnajb4. En xeral, a resposta ao choque térmico no transcriptoma hepático foi máis leve que a resposta in vitro, o que é probablemente consecuencia da activación de mecanismos compensatorios. Estes mecanismos inclúen o sistema neuroendocrino e teñen como consecuencia un aumento da protección dos tecidos e das capacidades termoxénicas, especialmente no peixe criado a temperaturas fluctuantes. Polo tanto, propoñemos que a cría de peixes a temperaturas fluctuantes debe ser incluida no conxunto de novas técnicas empegadas en peixes criados con fins de repoboamento, non obstante, outros protocolos de temperatura deben ser investigados. Dado que unha importante limitación para a avaliación de novas técnicas de cría é a falta dun xenoma de referencia para experimentos de mapeado do transcriptoma, no Artigo III temos ensamblado un xenoma de referencia para o esturión Atlántico, combinando tecnoloxías de secuenciación de curta e longa lectura. O xenoma do esturión Atlántico evidencia por primeira vez a presenza dun evento de duplicación específico de esturión (SR) e independente do peixe-espátula (Polyodon spathula). A presenza de xenes Hox duplicados, xunto con estudos filoxenéticos e de sintenia e os resultados da análise de loci microsatélite suxire que os esturións teñen unha orixe paleotetraploide e que a rediploidización é aínda un proceso activo. En resumo, os resultados presentados nesta tese avanzan no campo da investigación con esturións. Os nosos resultados suxiren que a cría de peixes baixo un réxime de temperatura fluctuante ten un efecto positivo durante un choque térmico subsecuente, pero a influencia de esta nova técnica de cría na supervivencia dos xuvenís tras a súa liberación no habitat a repoboar aínda debe ser avaliada. Contudo, suxerimos que o traballo no futuro ten que estar centrando na optimización dos métodos de cría en programas de repoboación e que o xenoma de referencia debe ser usado. The European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement no. 642893: Improved Production Strategies for Endangered Freshwater Species, “IMPRESS”.
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- 2020
120. Association between stress, metabolism, and growth in Ichthyophthirius multifiliis infected rainbow trout gills: Transcriptomic evidence.
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Syahputra, Khairul, Kania, Per W., Al-Jubury, Azmi, Marnis, Huria, Mathiessen, Heidi, Dirks, Ron P., and Buchmann, Kurt
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RAINBOW trout , *GILLS , *FISH physiology , *METABOLISM , *ENERGY metabolism , *TRANSCRIPTION factors - Abstract
Infection of rainbow trout with the parasitic ciliate Ichthyophthirius multifiliis induces a stress response which can be monitored by serum cortisol measurements reflecting involvement of the pituitary-interrenal cell axis in parasite infected fish. Molecular mechanisms associated with the stress response have been less well elucidated - especially with regard to stress effects on metabolism and growth in infected gills – which is the focus of the present study. A controlled experimental infection was performed followed by serum and gill sampling at day 8 post-infection. The stress reactions in the fish were elucidated by measurements of serum cortisol and lysozyme activity, and expression in gills of genes associated with stress, metabolism, and growth. Infection induced a marked elevation of serum cortisol and lysozyme levels, and major changes of metabolic processes in gills. A number of key genes involved in stress (hsps , foxo , saa) and energy metabolism (cyps , pfkfb3 , rrm2 , mthfd1l) were activated in response to infection whereas several central genes related to growth factors (wnt7a , igfbp7 , insr , fgf7 , tgfb3 , tgfbr1 , tgfbr3) were suppressed. A number of potential transcription factors including myc , e2f1 , stat1 , jun , e2f2 , e2f3 , and gli2 were found associated with regulation of genes in gills during infection. Mechanisms linking infection of mucosal surfaces and systemic responses are discussed. • Ichthyophthirius multifiliis induced a state of stress in rainbow trout. • A marked increase of serum cortisol and lysozyme was associated with the infection. • The parasite induced dramatic transcriptomic changes in infected gills. • Key genes associated with stress and energy metabolism were activated. • The majority of genes related to growth were down-regulated in infected gills. [ABSTRACT FROM AUTHOR]
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- 2020
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121. Author Correction: Rapid de novo assembly of the European eel genome from nanopore sequencing reads.
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Jansen, Hans J., Liem, Michael, Jong-Raadsen, Susanne A., Dufour, Sylvie, Weltzien, Finn-Arne, Swinkels, William, Koelewijn, Alex, Palstra, Arjan P., Pelster, Bernd, Spaink, Herman P., van den Thillart, Guido E., Dirks, Ron P., and Henkel, Christiaan V.
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A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper. [ABSTRACT FROM AUTHOR]
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- 2019
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122. Functional divergence of thyrotropin beta-subunit paralogs gives new insights into salmon smoltification metamorphosis.
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Fleming, Mitchell S., Maugars, Gersende, Lafont, Anne-Gaëlle, Rancon, Jocelyn, Fontaine, Romain, Nourizadeh-Lillabadi, Rasoul, Weltzien, Finn-Arne, Yebra-Pimentel, Elena Santidrian, Dirks, Ron, McCormick, Stephen D., Rousseau, Karine, Martin, Patrick, and Dufour, Sylvie
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Smoltification is a metamorphic event in salmon life history, which initiates downstream migration and pre-adapts juvenile salmon for seawater entry. While a number of reports concern thyroid hormones and smoltification, few and inconclusive studies have addressed the potential role of thyrotropin (TSH). TSH is composed of a α-subunit common to gonadotropins, and a β-subunit conferring hormone specificity. We report the presence and functional divergence of duplicated TSH β-subunit paralogs (tshβa and tshβb) in Atlantic salmon. Phylogeny and synteny analyses allowed us to infer that they originated from teleost-specific whole genome duplication. Expression profiles of both paralogs in the pituitary were measured by qPCR throughout smoltification in Atlantic salmon from the endangered Loire-Allier population raised in a conservation hatchery. This revealed a striking peak of tshβb expression in April, concomitant with downstream migration initiation, while tshβa expression remained relatively constant. In situ hybridization showed two distinct pituitary cell populations, tshβa cells in the anterior adenohypophysis, and tshβb cells near to the pituitary stalk, a location comparable to the pars tuberalis TSH cells involved in seasonal physiology and behaviour in birds and mammals. Functional divergence of tshβ paralogs in Atlantic salmon supports a specific role of tshβb in smoltification. [ABSTRACT FROM AUTHOR]
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- 2019
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123. A chromosome-scale genome assembly of Rauvolfia tetraphylla facilitates identification of the complete ajmaline biosynthetic pathway.
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Lezin E, Carqueijeiro I, Cuello C, Durand M, Jansen HJ, Vergès V, Birer Williams C, Oudin A, Dugé de Bernonville T, Petrignet J, Celton N, St-Pierre B, Papon N, Sun C, Dirks RP, O'Connor SE, Jensen MK, Besseau S, and Courdavault V
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- Ajmaline metabolism, Biosynthetic Pathways genetics, Rauwolfia genetics, Rauwolfia metabolism
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- 2024
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124. The Madagascar palm genome provides new insights on the evolution of Apocynaceae specialized metabolism.
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Cuello C, Jansen HJ, Abdallah C, Zamar Mbadinga DL, Birer Williams C, Durand M, Oudin A, Papon N, Giglioli-Guivarc'h N, Dirks RP, Jensen MK, O'Connor SE, Besseau S, and Courdavault V
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Specialized metabolites possess diverse interesting biological activities and some cardenolides- and monoterpene indole alkaloids- (MIAs) derived pharmaceuticals are currently used to treat human diseases such as cancers or hypertension. While these two families of biocompounds are produced by specific subfamilies of Apocynaceae , one member of this medicinal plant family, the succulent tree Pachypodium lamerei Drake (also known as Madagascar palm), does not produce such specialized metabolites. To explore the evolutionary paths that have led to the emergence and loss of cardenolide and MIA biosynthesis in Apocynaceae , we sequenced and assembled the P. lamerei genome by combining Oxford Nanopore Technologies long-reads and Illumina short-reads. Phylogenomics revealed that, among the Apocynaceae whose genomes have been sequenced, the Madagascar palm is so far the species closest to the common ancestor between MIA producers/non-MIA producers. Transposable elements, constituting 72.48% of the genome, emerge as potential key players in shaping genomic architecture and influencing specialized metabolic pathways. The absence of crucial MIA biosynthetic genes such as strictosidine synthase in P. lamerei and non- Rauvolfioideae species hints at a transposon-mediated mechanism behind gene loss. Phylogenetic analysis not only showcases the evolutionary divergence of specialized metabolite biosynthesis within Apocynaceae but also underscores the role of transposable elements in this intricate process. Moreover, we shed light on the low conservation of enzymes involved in the final stages of MIA biosynthesis in the distinct MIA-producing plant families, inferring independent gains of these specialized enzymes along the evolution of these medicinal plant clades. Overall, this study marks a leap forward in understanding the genomic dynamics underpinning the evolution of specialized metabolites biosynthesis in the Apocynaceae family, with transposons emerging as potential architects of genomics restructuring and gene loss., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:Vincent Courdavault reports financial support was provided by Horizon Europe. Michael Krogh Jensen reports a relationship with BioMIA that includes: board membership and employment. Ron Dirks reports a relationship with Future Genomics Technologies that includes: board membership and employment. Hans Jensen reports a relationship with Future Genomics Technologies that includes: board membership and employment. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)
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- 2024
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125. Development of Potent Mcl-1 Inhibitors: Structural Investigations on Macrocycles Originating from a DNA-Encoded Chemical Library Screen.
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Hekking KFW, Maroto S, van Kekem K, Haasjes FS, Slootweg JC, Oude Alink PGB, Dirks R, Sardana M, Bolster MG, Kuijpers B, Smith D, Doodeman R, Scheepstra M, Zech B, Mulvihill M, Renzetti LM, Babiss L, Centrella PA, Clark MA, Cuozzo JW, Guié MA, Sigel E, Habeshian S, Hupp CD, Liu J, Thomson HA, Zhang Y, Keefe AD, Müller G, and Gremmen S
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- Humans, Myeloid Cell Leukemia Sequence 1 Protein metabolism, Apoptosis, Molecular Conformation, DNA, Cell Line, Tumor, Proto-Oncogene Proteins c-bcl-2 metabolism, Neoplasms, Antineoplastic Agents pharmacology, Antineoplastic Agents chemistry
- Abstract
Evasion of apoptosis is critical for the development and growth of tumors. The pro-survival protein myeloid cell leukemia 1 (Mcl-1) is an antiapoptotic member of the Bcl-2 family, associated with tumor aggressiveness, poor survival, and drug resistance. Development of Mcl-1 inhibitors implies blocking of protein-protein interactions, generally requiring a lengthy optimization process of large, complex molecules. Herein, we describe the use of DNA-encoded chemical library synthesis and screening to directly generate complex, yet conformationally privileged macrocyclic hits that serve as Mcl-1 inhibitors. By applying a conceptual combination of conformational analysis and structure-based design in combination with a robust synthetic platform allowing rapid analoging, we optimized in vitro potency of a lead series into the low nanomolar regime. Additionally, we demonstrate fine-tuning of the physicochemical properties of the macrocyclic compounds, resulting in the identification of lead candidates 57 / 59 with a balanced profile, which are suitable for future development toward therapeutic use.
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- 2024
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126. The Rauvolfia tetraphylla genome suggests multiple distinct biosynthetic routes for yohimbane monoterpene indole alkaloids.
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Stander EA, Lehka B, Carqueijeiro I, Cuello C, Hansson FG, Jansen HJ, Dugé De Bernonville T, Birer Williams C, Vergès V, Lezin E, Lorensen MDBB, Dang TT, Oudin A, Lanoue A, Durand M, Giglioli-Guivarc'h N, Janfelt C, Papon N, Dirks RP, O'connor SE, Jensen MK, Besseau S, and Courdavault V
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- Monoterpenes, Indole Alkaloids metabolism, Rauwolfia genetics, Rauwolfia metabolism
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Monoterpene indole alkaloids (MIAs) are a structurally diverse family of specialized metabolites mainly produced in Gentianales to cope with environmental challenges. Due to their pharmacological properties, the biosynthetic modalities of several MIA types have been elucidated but not that of the yohimbanes. Here, we combine metabolomics, proteomics, transcriptomics and genome sequencing of Rauvolfia tetraphylla with machine learning to discover the unexpected multiple actors of this natural product synthesis. We identify a medium chain dehydrogenase/reductase (MDR) that produces a mixture of four diastereomers of yohimbanes including the well-known yohimbine and rauwolscine. In addition to this multifunctional yohimbane synthase (YOS), an MDR synthesizing mainly heteroyohimbanes and the short chain dehydrogenase vitrosamine synthase also display a yohimbane synthase side activity. Lastly, we establish that the combination of geissoschizine synthase with at least three other MDRs also produces a yohimbane mixture thus shedding light on the complex mechanisms evolved for the synthesis of these plant bioactives., (© 2023. The Author(s).)
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- 2023
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127. The induction of oocyte maturation and ovulation in the European eel ( Anguilla anguilla ): in vitro and in vivo comparison of progesterone with 17α,20β-dihydroxy-4-pregnen-3-one.
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Jéhannet P, Palstra AP, Meijerhof M, Schipper H, Giménez IN, Dirks RP, Swinkels W, Heinsbroek LTN, and Komen H
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Ovulation in European eel is induced by injection of 17α,20β-dihydroxy-4-pregnen-3-one (DHP) as the maturation-inducing hormone (MIH). Female eels need to ovulate within 18 h after injection to release good quality eggs. Progesterone (P), as an upstream precursor of DHP, may promote endogenous DHP production and improve egg quality. The purpose of this study was therefore to compare treatment of P with DHP on batch level, in vitro , to determine dose-response effects, and in vivo , at a single dose. For the in vitro experiment, ovarian tissue was extracted and placed in culture plates containing hormone-free medium and media supplemented with the treatment: DHP at 1, 10 and 100 ng mL
-1 , or P at 10, 100 and 1,000 ng mL-1 . At the start of incubation, the folliculated oocytes were sampled for histology, microscopy and qPCR. After incubation for 12 and 18 h, the oocytes were sampled for microscopy and qPCR analysis. For the in vivo experiment, females were either injected with DHP or P at a dose of 2 mg kg-1 to assess their effects on ovulation and reproductive success. At the moment of release, eggs were sampled for RNA sequencing to compare effects of DHP and P on the expression of genes involved in egg quality aspects. Remaining eggs were fertilized and larval viability was recorded. Both DHP and P were able to induce GVBD (DHP at 10 and 100 ng mL-1 , P at 100 and 1,000 ng mL-1 ) in vitro . Expression of genes involved in oocyte maturation and ovulation was similar in vitro for both DHP and P treatments. Regarding the in vivo results, RNAseq results reflected similar DHP and P effects on the expression of genes involved in egg quality aspects. Females injected with either DHP or P ovulated, released eggs, and were equally able to produce larvae without any differences in reproductive success. Our results support the conclusion that DHP and P work equally well in vitro and in vivo . P is more attractive to apply as the price is 3,000 times lower than the price of DHP., Competing Interests: Author IG was employed by the company Rara Avis. Author RD was employed by the company Future Genomics Technologies B.V. Author WS was employed by the company Palingkwekerij Koolen B.V. Author LH was employed by the company Wageningen Eel Reproduction Experts B.V. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Jéhannet, Palstra, Meijerhof, Schipper, Giménez, Dirks, Swinkels, Heinsbroek and Komen.)- Published
- 2023
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128. Genome structures resolve the early diversification of teleost fishes.
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Parey E, Louis A, Montfort J, Bouchez O, Roques C, Iampietro C, Lluch J, Castinel A, Donnadieu C, Desvignes T, Floi Bucao C, Jouanno E, Wen M, Mejri S, Dirks R, Jansen H, Henkel C, Chen WJ, Zahm M, Cabau C, Klopp C, Thompson AW, Robinson-Rechavi M, Braasch I, Lecointre G, Bobe J, Postlethwait JH, Berthelot C, Roest Crollius H, and Guiguen Y
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- Animals, Eels classification, Eels genetics, Genome, Phylogeny, Zebrafish classification, Zebrafish genetics, Biological Evolution, Fishes classification, Fishes genetics
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Accurate species phylogenies are a prerequisite for all evolutionary research. Teleosts are the largest and most diversified group of extant vertebrates, but relationships among their three oldest extant lineages remain unresolved. On the basis of seven high-quality new genome assemblies in Elopomorpha (tarpons, eels), we revisited the topology of the deepest branches of the teleost phylogeny using independent gene sequence and chromosomal rearrangement phylogenomic approaches. These analyses converged to a single scenario that unambiguously places the Elopomorpha and Osteoglossomorpha (arapaima, elephantnose fish) in a monophyletic sister group to all other teleosts, i.e., the Clupeocephala lineage (zebrafish, medaka). This finding resolves more than 50 years of controversy on the evolutionary relationships of these lineages and highlights the power of combining different levels of genome-wide information to solve complex phylogenies.
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- 2023
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129. The Vinca minor genome highlights conserved evolutionary traits in monoterpene indole alkaloid synthesis.
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Stander EA, Cuello C, Birer-Williams C, Kulagina N, Jansen HJ, Carqueijeiro I, Méteignier LV, Vergès V, Oudin A, Papon N, Dirks RP, Jensen MK, O'Connor SE, Dugé de Bernonville T, Besseau S, and Courdavault V
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- Monoterpenes, Phylogeny, Biological Evolution, Phenotype, Vinca
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Vinca minor, also known as the lesser periwinkle, is a well-known species from the Apocynaceae, native to central and southern Europe. This plant synthesizes monoterpene indole alkaloids, which are a class of specialized metabolites displaying a wide range of bioactive- and pharmacologically important properties. Within the almost 50 monoterpene indole alkaloids it produces, V. minor mainly accumulates vincamine, which is commercially used as a nootropic. Using a combination of Oxford Nanopore Technologies long read- and Illumina short-read sequencing, a 679,098 Mb V. minor genome was assembled into 296 scaffolds with an N50 scaffold length of 6 Mb, and encoding 29,624 genes. These genes were functionally annotated and used in a comparative genomic analysis to establish gene families and to investigate gene family expansion and contraction across the phylogenetic tree. Furthermore, homology-based monoterpene indole alkaloid gene predictions together with a metabolic analysis across 4 different V. minor tissue types guided the identification of candidate monoterpene indole alkaloid genes. These candidates were finally used to identify monoterpene indole alkaloid gene clusters, which combined with synteny analysis allowed for the discovery of a functionally validated vincadifformine-16-hydroxylase, reinforcing the potential of this dataset for monoterpene indole alkaloids gene discovery. It is expected that access to these resources will facilitate the elucidation of unknown monoterpene indole alkaloid biosynthetic routes with the potential of transferring these pathways to heterologous expression systems for large-scale monoterpene indole alkaloid production., (© The Author(s) 2022. Published by Oxford University Press on behalf of Genetics Society of America.)
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- 2022
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130. Expression of transport proteins in the rete mirabile of european silver and yellow eel.
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Schneebauer G, Drechsel V, Dirks R, Faserl K, Sarg B, and Pelster B
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- Air Sacs metabolism, Animals, Biological Transport, Carrier Proteins metabolism, Lactic Acid metabolism, Anguilla genetics, Eels genetics
- Abstract
Background: In physoclist fishes filling of the swimbladder requires acid secretion of gas gland cells to switch on the Root effect and subsequent countercurrent concentration of the initial gas partial pressure increase by back-diffusion of gas molecules in the rete mirabile. It is generally assumed that the rete mirabile functions as a passive exchanger, but a detailed analysis of lactate and water movements in the rete mirabile of the eel revealed that lactate is diffusing back in the rete. In the present study we therefore test the hypothesis that expression of transport proteins in rete capillaries allows for back-diffusion of ions and metabolites, which would support the countercurrent concentrating capacity of the rete mirabile. It is also assumed that in silver eels, the migratory stage of the eel, the expression of transport proteins would be enhanced., Results: Analysis of the transcriptome and of the proteome of rete mirabile tissue of the European eel revealed the expression of a large number of membrane ion and metabolite transport proteins, including monocarboxylate and glucose transport proteins. In addition, ion channel proteins, Ca
2+ -ATPase, Na+ /K+ -ATPase and also F1 F0 -ATP synthase were detected. In contrast to our expectation in silver eels the expression of these transport proteins was not elevated as compared to yellow eels. A remarkable number of enzymes degrading reactive oxygen species (ROS) was detected in rete capillaries., Conclusions: Our results reveal the expression of a large number of transport proteins in rete capillaries, so that the back diffusion of ions and metabolites, in particular lactate, may significantly enhance the countercurrent concentrating ability of the rete. Metabolic pathways allowing for aerobic generation of ATP supporting secondary active transport mechanisms are established. Rete tissue appears to be equipped with a high ROS defense capacity, preventing damage of the tissue due to the high oxygen partial pressures generated in the countercurrent system., (© 2021. The Author(s).)- Published
- 2021
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131. Genome and transcriptome analysis of the beet armyworm Spodoptera exigua reveals targets for pest control.
- Author
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Simon S, Breeschoten T, Jansen HJ, Dirks RP, Schranz ME, and Ros VID
- Subjects
- Animals, Female, Gene Expression Profiling, Larva, Pest Control, Pupa, Spodoptera genetics, Beta vulgaris
- Abstract
The genus Spodoptera (Lepidoptera: Noctuidae) includes some of the most infamous insect pests of cultivated plants including Spodoptera frugiperda, Spodoptera litura, and Spodoptera exigua. To effectively develop targeted pest control strategies for diverse Spodoptera species, genomic resources are highly desired. To this aim, we provide the genome assembly and developmental transcriptome comprising all major life stages of S. exigua, the beet armyworm. Spodoptera exigua is a polyphagous herbivore that can feed on > 130 host plants, including several economically important crops. The 419 Mb beet armyworm genome was sequenced from a female S. exigua pupa. Using a hybrid genome sequencing approach (Nanopore long-read data and Illumina short read), a high-quality genome assembly was achieved (N50 = 1.1 Mb). An official gene set (18,477 transcripts) was generated by automatic annotation and by using transcriptomic RNA-seq datasets of 18 S. exigua samples as supporting evidence. In-depth analyses of developmental stage-specific expression combined with gene tree analyses of identified homologous genes across Lepidoptera genomes revealed four potential genes of interest (three of them Spodoptera-specific) upregulated during first- and third-instar larval stages for targeted pest-outbreak management. The beet armyworm genome sequence and developmental transcriptome covering all major developmental stages provide critical insights into the biology of this devastating polyphagous insect pest species worldwide. In addition, comparative genomic analyses across Lepidoptera significantly advance our knowledge to further control other invasive Spodoptera species and reveals potential lineage-specific target genes for pest control strategies., (© The Author(s) 2021. Published by Oxford University Press on behalf of Genetics Society of America.)
- Published
- 2021
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132. Innate immune-gene expression during experimental amyloodiniosis in European seabass (Dicentrarchus labrax).
- Author
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Byadgi O, Massimo M, Dirks RP, Pallavicini A, Bron JE, Ireland JH, Volpatti D, Galeotti M, and Beraldo P
- Subjects
- Animals, Gills parasitology, Head Kidney immunology, Head Kidney parasitology, Immunity, Innate immunology, RNA, Messenger genetics, Bass immunology, Dinoflagellida immunology, Fish Diseases immunology, Gene Expression, Immunity, Innate genetics, Protozoan Infections, Animal immunology
- Abstract
The ectoparasite protozoan Amyloodinium ocellatum (AO) is the causative agent of amyloodiniosis in European seabass (ESB, Dicentrarchus labrax). There is a lack of information about basic molecular immune response mechanisms of ESB during AO infestation. Therefore, to compare gene expression between experimental AO-infested ESB tissues and uninfested ESB tissues (gills and head kidney) RNA-seq was adopted. The RNA-seq revealed multiple differentially expressed genes (DEG), namely 679 upregulated genes and 360 downregulated genes in the gills, and 206 upregulated genes and 170 downregulated genes in head kidney. In gills, genes related to the immune system (perforin, CC1) and protein binding were upregulated. Several genes involved in IFN related pathways were upregulated in the head kidney. Subsequently, to validate the DEG from amyloodiniosis, 26 ESB (mean weight 14 g) per tank in triplicate were bath challenged for 2 h with AO (3.5 × 10
6 /tank; 70 dinospores/mL) under controlled conditions (26-28 °C and 34‰ salinity). As a control group (non-infested), 26 ESB per tank in triplicate were also used. Changes in the expression of innate immune genes in gills and head kidney at 2, 3, 5, 7 and 23 dpi were analysed using real-time PCR. The results indicated that the expression of cytokines (CC1, IL-8) and antimicrobial peptide (Hep) were strongly stimulated and reached a peak at 5 dpi in the early infestation stage, followed by a gradual reduction in the recovery stage (23 dpi). Noticeably, the immunoglobulin (IgM) expression was higher at 23 dpi compared to 7 dpi. Furthermore, in-situ hybridization showed positive signals of CC1 mRNA in AO infested gills compared to the control group. Altogether, chemokines were involved in the immune process under AO infestation and this evidence allows a better understanding of the immune response in European seabass during amyloodiniosis., (Copyright © 2021. Published by Elsevier B.V.)- Published
- 2021
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133. Temperature training improves transcriptional homeostasis after heat shock in juvenile Atlantic sturgeon (Acipenser oxyrinchus).
- Author
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Yebra-Pimentel ES, Reis B, Gessner J, Wuertz S, and Dirks RPH
- Subjects
- Animals, Adaptation, Physiological, Fishes physiology, Gene Expression Regulation physiology, Heat-Shock Response, Homeostasis
- Abstract
Exposure to high temperatures can lead to thermotolerance in fish, which is hypothesized to potentially improve post-release survival in species under restocking programs, like Atlantic sturgeon. The aim of this study was to determine whether Atlantic sturgeon juveniles exposed to a 4-week temperature treatment respond differently to a subsequent heat shock than juveniles exposed to heat shock for the first time (naive fish). Response to heat shock was assessed by mapping the liver transcriptome. In total, 838 unique contigs were differentially expressed between the trained and the control group (592 downregulated, 261 upregulated, and 15 down- or upregulated, depending on the condition), corresponding to genes involved in the response to heat, tissue damage, proteolysis, and metabolism. Temperature-trained fish showed 2-4-fold fewer dysregulated contigs than naive fish, indicating their ability to maintain and recover homeostasis faster. During heat shock, hspc1 was upregulated in both experimental groups, while hspa1 and dnaja4 were exclusively upregulated in the control. Overall, compensatory mechanisms were observed in addition to the heat shock response. Only two genes, fgg and apnl, were upregulated at nearly all timepoints in both groups. Peptidases were more strongly downregulated in control fish, which also showed a reduction in lipid metabolism during recovery. Keratins, pck1, gadd45ga, and gadd45gb were differentially expressed between trained and control fish, and due to their roles in tissue protection and ER stress reduction, they might be responsible for the maintenance of the transcriptional homeostasis observed in trained fish.
- Published
- 2020
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134. Application of Caenorhabditis elegans (nematode) and Danio rerio embryo (zebrafish) as model systems to screen for developmental and reproductive toxicity of Piperazine compounds.
- Author
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Racz PI, Wildwater M, Rooseboom M, Kerkhof E, Pieters R, Yebra-Pimentel ES, Dirks RP, Spaink HP, Smulders C, and Whale GF
- Subjects
- Animals, Caenorhabditis elegans embryology, Embryo, Nonmammalian, Models, Animal, Caenorhabditis elegans drug effects, Embryonic Development drug effects, Piperazines toxicity, Reproduction drug effects, Zebrafish embryology
- Abstract
To enable selection of novel chemicals for new processes, there is a recognized need for alternative toxicity screening assays to assess potential risks to man and the environment. For human health hazard assessment these screening assays need to be translational to humans, have high throughput capability, and from an animal welfare perspective be harmonized with the principles of the 3Rs (Reduction, Refinement, Replacement). In the area of toxicology a number of cell culture systems are available but while these have some predictive value, they are not ideally suited for the prediction of developmental and reproductive toxicology (DART). This is because they often lack biotransformation capacity, multicellular or multi- organ complexity, for example, the hypothalamus pituitary gonad (HPG) axis and the complete life cycle of whole organisms. To try to overcome some of these limitations in this study, we have used Caenorhabditis elegans (nematode) and Danio rerio embryos (zebrafish) as alternative assays for DART hazard assessment of some candidate chemicals being considered for a new commercial application. Nematodes exposed to Piperazine and one of the analogs tested showed a slight delay in development compared to untreated animals but only at high concentrations and with Piperazine as the most sensitive compound. Total brood size of the nematodes was also reduced primarily by Piperazine and one of the analogs. In zebrafish Piperazine and analogs showed developmental delays. Malformations and mortality in individual fish were also scored. Significant malformations were most sensitively identified with Piperazine, significant mortality was only observed in Piperazine and only at the higest dose. Thus, Piperazine seemed the most toxic compound for both nematodes and zebrafish. The results of the nematode and zebrafish studies were in alignment with data obtained from conventional mammalian toxicity studies indicating that these have potential as developmental toxicity screening systems. The results of these studies also provided reassurance that none of the Piperazines tested are likely to have any significant developmental and/or reproductive toxicity issues to humans when used in their commercial applications., (Copyright © 2017. Published by Elsevier Ltd.)
- Published
- 2017
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135. De novo whole-genome assembly of a wild type yeast isolate using nanopore sequencing.
- Author
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Liem M, Jansen HJ, Dirks RP, Henkel CV, van Heusden GPH, Lemmers RJLF, Omer T, Shao S, Punt PJ, and Spaink HP
- Abstract
Background : The introduction of the MinION sequencing device by Oxford Nanopore Technologies may greatly accelerate whole genome sequencing. Nanopore sequence data offers great potential for de novo assembly of complex genomes without using other technologies. Furthermore, Nanopore data combined with other sequencing technologies is highly useful for accurate annotation of all genes in the genome. In this manuscript we used nanopore sequencing as a tool to classify yeast strains. Methods : We compared various technical and software developments for the nanopore sequencing protocol, showing that the R9 chemistry is, as predicted, higher in quality than R7.3 chemistry. The R9 chemistry is an essential improvement for assembly of the extremely AT-rich mitochondrial genome. We double corrected assemblies from four different assemblers with PILON and assessed sequence correctness before and after PILON correction with a set of 290 Fungi genes using BUSCO. Results : In this study, we used this new technology to sequence and de novo assemble the genome of a recently isolated ethanologenic yeast strain, and compared the results with those obtained by classical Illumina short read sequencing. This strain was originally named Candida vartiovaarae ( Torulopsis vartiovaarae ) based on ribosomal RNA sequencing. We show that the assembly using nanopore data is much more contiguous than the assembly using short read data. We also compared various technical and software developments for the nanopore sequencing protocol, showing that nanopore-derived assemblies provide the highest contiguity. Conclusions : The mitochondrial and chromosomal genome sequences showed that our strain is clearly distinct from other yeast taxons and most closely related to published Cyberlindnera species. In conclusion, MinION-mediated long read sequencing can be used for high quality de novo assembly of new eukaryotic microbial genomes., Competing Interests: Competing interests: HJJ and CVH are members of the Nanopore Community, and have previously received flow cells free of charge, as well as travel expense reimbursements from Oxford Nanopore Technologies.
- Published
- 2017
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136. The king cobra genome reveals dynamic gene evolution and adaptation in the snake venom system.
- Author
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Vonk FJ, Casewell NR, Henkel CV, Heimberg AM, Jansen HJ, McCleary RJ, Kerkkamp HM, Vos RA, Guerreiro I, Calvete JJ, Wüster W, Woods AE, Logan JM, Harrison RA, Castoe TA, de Koning AP, Pollock DD, Yandell M, Calderon D, Renjifo C, Currier RB, Salgado D, Pla D, Sanz L, Hyder AS, Ribeiro JM, Arntzen JW, van den Thillart GE, Boetzer M, Pirovano W, Dirks RP, Spaink HP, Duboule D, McGlinn E, Kini RM, and Richardson MK
- Subjects
- Animals, Exocrine Glands metabolism, MicroRNAs genetics, MicroRNAs metabolism, Adaptation, Biological physiology, Elapid Venoms genetics, Elapid Venoms metabolism, Elapidae genetics, Elapidae metabolism, Evolution, Molecular, Genome physiology, Transcriptome physiology
- Abstract
Snakes are limbless predators, and many species use venom to help overpower relatively large, agile prey. Snake venoms are complex protein mixtures encoded by several multilocus gene families that function synergistically to cause incapacitation. To examine venom evolution, we sequenced and interrogated the genome of a venomous snake, the king cobra (Ophiophagus hannah), and compared it, together with our unique transcriptome, microRNA, and proteome datasets from this species, with data from other vertebrates. In contrast to the platypus, the only other venomous vertebrate with a sequenced genome, we find that snake toxin genes evolve through several distinct co-option mechanisms and exhibit surprisingly variable levels of gene duplication and directional selection that correlate with their functional importance in prey capture. The enigmatic accessory venom gland shows a very different pattern of toxin gene expression from the main venom gland and seems to have recruited toxin-like lectin genes repeatedly for new nontoxic functions. In addition, tissue-specific microRNA analyses suggested the co-option of core genetic regulatory components of the venom secretory system from a pancreatic origin. Although the king cobra is limbless, we recovered coding sequences for all Hox genes involved in amniote limb development, with the exception of Hoxd12. Our results provide a unique view of the origin and evolution of snake venom and reveal multiple genome-level adaptive responses to natural selection in this complex biological weapon system. More generally, they provide insight into mechanisms of protein evolution under strong selection.
- Published
- 2013
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137. Urochordate betagamma-crystallin and the evolutionary origin of the vertebrate eye lens.
- Author
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Shimeld SM, Purkiss AG, Dirks RP, Bateman OA, Slingsby C, and Lubsen NH
- Subjects
- Amino Acid Sequence, Animals, Ciona intestinalis anatomy & histology, Cloning, Molecular, Crystallization, Gene Expression Regulation genetics, Green Fluorescent Proteins, Likelihood Functions, Models, Genetic, Molecular Sequence Data, Sequence Alignment, X-Ray Diffraction, Xenopus, beta-Crystallins chemistry, gamma-Crystallins chemistry, Ciona intestinalis genetics, Evolution, Molecular, Lens, Crystalline anatomy & histology, Models, Molecular, Phylogeny, beta-Crystallins genetics, gamma-Crystallins genetics
- Abstract
A refracting lens is a key component of our image-forming camera eye; however, its evolutionary origin is unknown because precursor structures appear absent in nonvertebrates. The vertebrate betagamma-crystallin genes encode abundant structural proteins critical for the function of the lens. We show that the urochordate Ciona intestinalis, which split from the vertebrate lineage before the evolution of the lens, has a single gene coding for a single domain monomeric betagamma-crystallin. The crystal structure of Ciona betagamma-crystallin is very similar to that of a vertebrate betagamma-crystallin domain, except for paired, occupied calcium binding sites. The Ciona betagamma-crystallin is only expressed in the palps and in the otolith, the pigmented sister cell of the light-sensing ocellus. The Ciona betagamma-crystallin promoter region targeted expression to the visual system, including lens, in transgenic Xenopus tadpoles. We conclude that the vertebrate betagamma-crystallins evolved from a single domain protein already expressed in the neuroectoderm of the prevertebrate ancestor. The conservation of the regulatory hierarchy controlling betagamma-crystallin expression between organisms with and without a lens shows that the evolutionary origin of the lens was based on co-option of pre-existing regulatory circuits controlling the expression of a key structural gene in a primitive light-sensing system.
- Published
- 2005
- Full Text
- View/download PDF
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