Unknown,Transcriptomics,Genomics,Proteomics

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Dam selectivity upon glass eel migration: insights from genes expression analysis.

ABSTRACT: Gene expression analyses have been performed on brain, liver and muscle of eels (Anguilla anguilla) sampled in three different segment of the Canal des Etangs ( artificial canal linking Arcachon Basin and Lacanau Lake). The present study relies on intraspecific differences of glass eels encountered below and above the water dams. Eels were collected using electric fishing under similar climatic and hydrological conditions. Individuals were sampled below the obstacle, close to the fishway entry in segment 1 (Pas du Bouc; +44° 50' 27.95, -1° 9' 8.09) and 2 (Langouarde, +44° 51' 32.92, -1° 9' 5.03). Fish from the segment 3 (Joncru; +44° 52' 57.13, -1° 8' 11.70) were sampled on the fishway, as water depth before the obstacle, approximately 2 meters, precluded the use of electro-fishing technique. Ten individuals were selected from each site according to their body size (between 83 and 155 mm) and health status (no externally visible pathogens) criteria. Internal parasites such as Anguilicola crassus were taken into account. Sampled and selected fish were immediately sacrified by decapitation and the whole brain, liver and muscle tissue (posterior bottom body part) were dissected and stored at -80°C in separate tubes with RNAlater buffer (Quiagen) for gene expression analysis. Microarray analysis was conducted using an European eel-specific array consisting of a total of 14,913 probes based on a large collection of high-throughput transcriptomic sequences (Pujolar et al. 2012). Probe sequences and further details on the microarray platform can be found on the GEO database under accession number GPL15124. A comparative analysis of gene expression was conducted between eels sampled in differents segments of Canal des Etangs. Total RNAs were extracted from the whole liver,muscle and brain using the Absolutely RNA RT-PCR Miniprep kit (Agilent) according to the manufacturer’s instructions. Three independent RNA pools, each consisting of three individuals RNA samples, were prepared for each sampling site and tissues. Due to technical problems, it was no possible to perform microarray analyses in muscle of eels sampled in forebay 3. Accordingly, DNA microarray analysis has been performed in a total of 24 pools. RNAs’ quality was evaluated by electrophoresis on a 1% agarose gel and their concentrations were determined by spectrophotometry. Total RNA was stored in RNAse free water at -80° C. Sample labelling and hybridization were conducted following the details in Pujolar et al. (2012). Hybridized slides were scanned at 5 µm resolution using an Agilent DNA microarray scanner. Slides were scanned at two different sensitivity levels (XDR Hi 100% and XDR Lo 10%) and the two linked images generated were analysed together. Data were extracted and background subtracted using the standard procedure in Agilent Feature Extraction (FE) software v. 9.5.1. Data were normalized separately for each tissue using a quantile normalization procedure using R (http://www.r-project.org).

ORGANISM(S): Anguilla anguilla

SUBMITTER: Massimo Milan

PROVIDER: E-GEOD-56040 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:In spite of many decades of research, the spawning migration of the European eel Anguilla anguilla from the European coast to the Sargasso Sea remains a mystery. In particular, the role of the swimbladder as a buoyancy regulating structure is not yet understood. In this study, we exercised silver eels in a swim tunnel under elevated hydrostatic pressure. The transcriptome of gas gland tissue of these exercised eels was then compared to the known transcriptome of not exercised (control) silver eel gas gland cells. Due to the high infection rate of the eel population with the swimbladder parasite Anguillicola crassus, the comparison also included an exercised group of silver eels with a heavily damaged swimbladder, and we compared the previously published transcriptome of not exercised silver eels with a highly damaged swimbladder with the exercised group of silver eels with a heavily damaged swimbladder. The comparisons of unexercised (control) silver eels with exercised silver eels with functional swimbladder (EF), as well as with exercised silver eels with damaged swimbladder(ED), both showed a significant elevation in transcripts related to glycolytic enzymes. This could also be observed within the comparison of unexercised silver eels with a highly infected swimbladder with exercised eels with a damaged swimbladder (DED). In contrast to EF, in ED a significant elevation in transcript numbers of mitochondrial NADH dehydrogenase was observed. While in EF the transcriptional changes suggested that acid production and secretion was enhanced, in ED these changes appeared to be related to thickened tissue and thus elevated diffusion distances. The remarkable number of differentially expressed transcripts coding for proteins connected to cAMP-dependent signaling pathways indicated that metabolic control in gas gland cells includes cAMP-dependent pathways. In contrast to ED, in EF significant transcriptional changes could be related to the reconstruction of the extracellular matrix, while in ED tissue repair and inflammation was more pronounced. Surprisingly, in exercised eels hypoxia inducible transcription factor expression was elevated. In EF, a large number of genes related to the circadian clock were transcriptionally modified, which may be connected to the circadian vertical migrations observed during the spawning migration.

Project description:Ecotoxicogenomic studies face the problem of the lack of gene sequence information available for toxicologically relevant non-model pollution sentinel species. In this sense, next generation sequencing technologies allow obtaining deep de novo sequence information cost-effectively. We have thus employed the platform GS-FLX of Roche technologies, in order to sequence the multitissue transcriptome of a fish species under severe population decline, the European eel (Anguilla anguilla), using normalized cDNA (Evrogen). The European eel could be successfully employed in pollution biomonitoring studies as it is a euryhaline teleost that is quite resistant to chemical exposure. We have completed a one whole plate sequencing-run using the Titanium kit of Roche from which a total of 7.8x105 reads were obtained with a length average of 303.53 bp. Annotated genes, for instance, could be further assessed as biomarkers of exposure to specific chemical compounds in marine, estuarine and river waters. Designed tool will be thus useful, in the study of: a.- the mode of action of chemical compounds in active monitoring studies using caged eels, b.- the physiological processes that are specific to freshwater eels such as the one of sexual development and reproduction, c.- reasons that could explain the disappearance of the species from European waters. In the first context, a caging experiment was performed to measure the putative effects of a paper industry effluent on eel hepatic transcription levels. For this purpose, eels were caged upstream and downstream to the SMURFFIT-KAPPA paper industry in Iurreta (Biscay, Basque Country). Thus, hepatic transcription profiles reflect the stress levels suffered by eels. The experiment contains a time serial sampling: T0 = before sampling; T1 = after 3 days of exposure; T2=after 15 days of exposure. In addition, caged eels were placed in 2 sites, Upstream (Up) and Downstream (Do) of the hotspot. 6 samples were dissected per sampling group (5 groups= ToUP; T1UP; T1Do; T2Up; T2Do), 30 samples in total

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Dam selectivity upon glass eel migration: insights from genes expression analysis.

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