123 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
- Author
-
Dirks, Ron P.H., primary, Klok, Erik Jan, additional, van Genesen, Siebe T., additional, Schoenmakers, John G.G., additional, and Lubsen, Nicolette H., additional
- Published
- 1996
- Full Text
- View/download PDF
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
- Author
-
Dirks, Ron P.H., primary, Onnekink, Carla, additional, Jansen, Hans J., additional, de Jong, Aard, additional, and Bloemers, Henri P.J., additional
- Published
- 1995
- Full Text
- View/download PDF
104. In vivofootprinting and functional analysis of the human c-sis/PDGF B gene promoter provides evidence for two binding sites for transcriptional activators
- Author
-
Dirks, Ron P. H., primary, Jansen, Hans J., additional, van Gerven, Bart, additional, Onnekink, Carla, additional, and Bloemers, Henri P. J., additional
- Published
- 1995
- Full Text
- View/download PDF
105. Testing Tuberculosis Drug Efficacy in a Zebrafish High-Throughput Translational Medicine Screen
- Author
-
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.
- Published
- 2014
- Full Text
- View/download PDF
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
- Author
-
DIRKS, Ron P. H., primary, JANSEN, Hans J., additional, ONNEKINK, Carla, additional, JONGE, Rob J. A., additional, and BLOEMERS, Henri P. J., additional
- Published
- 1993
- Full Text
- View/download PDF
107. Localization and functional analysis of DNase-I-hypersensitive sites in the human c-sis/PDGF-B gene transcription unit and its flanking regions
- Author
-
DIRKS, Ron P. H., primary, JANSEN, Hans J., additional, GERRITSMA, Jort, additional, ONNEKINK, Carla, additional, and BLOEMERS, Henri P. J., additional
- Published
- 1993
- Full Text
- View/download PDF
108. Splicing of the platelet-derived-growth-factor A-chain mRNA in human malignant mesothelioma cell lines and regulation of its expression
- Author
-
LANGERAK, Anthonie W., primary, DIRKS, Ron P. H., additional, and VERSNEL, Marjan A., additional
- Published
- 1992
- Full Text
- View/download PDF
109. Cell-Autonomous TrkB Signaling in Presynaptic Retinal Ganglion Cells Mediates Axon Arbor Growth and Synapse Maturation during the Establishment of Retinotectal Synaptic Connectivity. .
- Author
-
Marshak, Sonya, Nikolakopoulou, Angeliki Maria, Dirks, Ron, Martens, Gerard J., and Cohen-Cory, Susana
- Subjects
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]
- Published
- 2007
- Full Text
- View/download PDF
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.
- Author
-
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.
- Subjects
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]
- Published
- 2004
- Full Text
- View/download PDF
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.
- Author
-
Dirks, Ron P. H., Kraft, Harry J., van Genesen, Siebe T., Klok, Erik J., Pfundt, Rolph, Schoenmakers, John G. G., and Lubsen, Nicolette H.
- Subjects
- *
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]
- Published
- 1996
- Full Text
- View/download PDF
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.
- Author
-
Dirks, Ron P. H., Jansen, Hans J., Onnekink, Carla, de Jonge, Rob J. A., and Bloemers, Henri P. J.
- Subjects
- *
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]
- Published
- 1993
- Full Text
- View/download PDF
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.
- Author
-
Dirks, Ron P.H., Onnekink, Carla, Jansen, Hans J., de Jong, Aard, and Bloemers, Henri P.J.
- Published
- 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.
- Author
-
Dirks, Ron P. H., Jansen, Hans J., van Gerven, Bart, Onnekink, Carla, and Bloemers, Henri P. J.
- Published
- 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
- Author
-
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)’.
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
- Author
-
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)’.
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
- Author
-
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
- Author
-
Wesseling, John G., primary, Dirks, Ron, additional, Smits, Man A., additional, and Schoenmakers, John G.G., additional
- Published
- 1989
- Full Text
- View/download PDF
119. New genomics and transcriptomics tools toward improving conservation strategies for sturgeons
- Author
-
Santidrian Yebra-Pimentel, Elena Maria, Dirks, Ron P., Weltzien, Finn-Arne, and Dufour, Sylvie
- Subjects
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”.
- Published
- 2020
120. The Vinca minor genome highlights conserved evolutionary traits in monoterpene indole alkaloid synthesis.
- Author
-
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
- Subjects
- Monoterpenes, Phylogeny, Biological Evolution, Phenotype, Vinca
- Abstract
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.)
- Published
- 2022
- Full Text
- View/download PDF
121. Genome and transcriptome analysis of the beet armyworm Spodoptera exigua reveals targets for pest control.
- Author
-
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
- Full Text
- View/download PDF
122. De novo whole-genome assembly of a wild type yeast isolate using nanopore sequencing.
- Author
-
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
- Full Text
- View/download PDF
123. Urochordate betagamma-crystallin and the evolutionary origin of the vertebrate eye lens.
- Author
-
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
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.