Gill transcriptome response to changes in environmental calcium in the green spotted puffer fish

Abstract Background Calcium ion is tightly regulated in body fluids and for euryhaline fish, which are exposed to rapid changes in environmental [Ca2+], homeostasis is especially challenging. The gill is the main organ of active calcium uptake and therefore plays a crucial role in the maintenance of calcium ion homeostasis. To study the molecular basis of the short-term responses to changing calcium availability, the whole gill transcriptome obtained by Super Serial Analysis of Gene Expression (SuperSAGE) of the euryhaline teleost green spotted puffer fish, Tetraodon nigroviridis, exposed to water with altered [Ca2+] was analysed. Results Transfer of T. nigroviridis from 10 ppt water salinity containing 2.9 mM Ca2+ to high (10 mM Ca2+ ) and low (0.01 mM Ca2+) calcium water of similar salinity for 2-12 h resulted in 1,339 differentially expressed SuperSAGE tags (26-bp transcript identifiers) in gills. Of these 869 tags (65%) were mapped to T. nigroviridis cDNAs or genomic DNA and 497 (57%) were assigned to known proteins. Thirteen percent of the genes matched multiple tags indicating alternative RNA transcripts. The main enriched gene ontology groups belong to Ca2+ signaling/homeostasis but also muscle contraction, cytoskeleton, energy production/homeostasis and tissue remodeling. K-means clustering identified co-expressed transcripts with distinct patterns in response to water [Ca2+] and exposure time. Conclusions The generated transcript expression patterns provide a framework of novel water calcium-responsive genes in the gill during the initial response after transfer to different [Ca2+]. This molecular response entails initial perception of alterations, activation of signaling networks and effectors and suggests active remodeling of cytoskeletal proteins during the initial acclimation process. Genes related to energy production and energy homeostasis are also up-regulated, probably reflecting the increased energetic needs of the acclimation response. This study is the first genome-wide transcriptome analysis of fish gills and is an important resource for future research on the short-term mechanisms involved in the gill acclimation responses to environmental Ca2+ changes and osmoregulation.Peer Reviewe

A transcriptome resource for the copepod Calanus glacialis across a range of culture temperatures

The copepod Calanus glacialis plays a key role in the Arctic pelagic ecosystem. Despite its ecological importance and ongoing climate changes, limited knowledge at the genomic level has hindered the understanding of the molecular processes underlying environmental stress responses and ecological adaptation. Transcriptome data was generated from an experiment with C glacialis copepodite (CV) subjected to five different temperatures. We obtained a total of 512,352 high-quality 454 pyrosequencing reads, which were assembled into 55,562 contigs distributed in 128 KEGG pathways. Functional analysis revealed numerous genes related to diverse biological functions and processes, including members of all major conserved signaling pathways. Comparative analysis of acclimated individuals to experimental temperatures has provided information about gene variations observed in several pathways (e.g. genes involved in energy, lipid and amino acid metabolism were shown to be down-regulated with increasing temperatures). These mRNA sequence resources will facilitate further studies on genomics and physiology-driven molecular processes in C glacialis and related species. (C) 2015 Elsevier B.V. All rights reserved.European Commission [FP7-ENV-2008-1-226248]; Polish National Science Centre [2011/03/B/NZ8/02876]; Portuguese Science and Technology Foundation (FCT) [FCT PTDC/MAR/72630]; FCT [SFRH/BPD/64174/2009]info:eu-repo/semantics/publishedVersio

Molecular cloning and sequence of Sparus aurata skeletal myosin light chains expressed in white muscle: developmental expression and thyroid regulation

Two full-length cDNA clones encoding the skeletal myosin light chain 2 (MLC2; 1452 bp) and myosin light chain 3 (MLC3; 972 bp) were isolated from a cDNA library prepared from gilthead sea bream Sparus aurata larvae. The MLC2 cDNA encoded a predicted protein of 170 residues that was 79 % identical to rabbit MLC2 over the entire length and 87 % identical within the Ca2+- binding region. The deduced amino acid sequence of MLC3 was 153 residues in length and was 91 % and 69% identical to the zebrafish and rabbit MLC3, respectively. Northern blot analysis revealed that in adults both transcripts were expressed in fast white muscle only. MLC2 appeared earlier in development: MLC2 transcripts were detectable from the beginning of segmentation, whereas MLC3 transcripts did not appear until 27 h post-fertilisation. At this developmental stage, a second MLC2 transcript of 0.89 kilobase-pairs was present. MLCs exhibited a different age-related pattern of response to varied thyroidal states, which were experimentally induced by the administration of 1 mgg-1 body mass of thyroxine (T4) or triiodothyronine (T3), or 5 ng g-1 body mass of the hypothyroidal compound thiourea; MLC3 expression was not significantly affected, whereas levels of MLC2 transcripts were significantly elevated in the white muscle only of juvenile sea bream after administration of T4. Although the mechanism of thyroidal regulation of MLC expression remains unknown, the present results suggest that different regulatory mechanisms exist for different MLCs

Free and conjugated androgen and progestin levels in the serum of stellate sturgeon (Acipenser stellatus Pallas) males treated with female coelomic fluid

Based on the supposition that female coelomic fluid contents have a certain chemical influence on sturgeons, blood serum concentrations of free and conjugated testosterone (T), 11-ketotestosterone (11KT), 17,20b,21-trihydroxy-4-pregnen- 3-one (20bS), 17,20b-dihydroxy-4-pregnen-3-one (DHP), 11-desoxycortisol (S) and free progesterone (P4) have been measured by radioimmunoassay (RIA) and enzyme-linked immuno-sorbent assay (ELISA) in stellate sturgeon (Acipenser stellatus Pallas) males treated with female coelomic fluid (CF); CF steroid levels have also been analyzed. After treatment a significant elevation of free 20bS and glucuronidated DHP and S levels and the decrease of free T and KT levels have been observed. The obtained data support the idea that the components of CF may play some part in pheromonal effects for sturgeon reproduction

The European sea bass <i>Dicentrarchus labrax</i> genome puzzle: comparative BAC-mapping and low coverage shotgun sequencing

BackgroundFood supply from the ocean is constrained by the shortage of domesticated and selected fish. Development of genomic models of economically important fishes should assist with the removal of this bottleneck. European sea bass Dicentrarchus labrax L. (Moronidae, Perciformes, Teleostei) is one of the most important fishes in European marine aquaculture; growing genomic resources put it on its way to serve as an economic model.ResultsEnd sequencing of a sea bass genomic BAC-library enabled the comparative mapping of the sea bass genome using the three-spined stickleback Gasterosteus aculeatus genome as a reference. BAC-end sequences (102,690) were aligned to the stickleback genome. The number of mappable BACs was improved using a two-fold coverage WGS dataset of sea bass resulting in a comparative BAC-map covering 87% of stickleback chromosomes with 588 BAC-contigs. The minimum size of 83 contigs covering 50% of the reference was 1.2 Mbp; the largest BAC-contig comprised 8.86 Mbp. More than 22,000 BAC-clones aligned with both ends to the reference genome. Intra-chromosomal rearrangements between sea bass and stickleback were identified. Size distributions of mapped BACs were used to calculate that the genome of sea bass may be only 1.3 fold larger than the 460 Mbp stickleback genome.ConclusionsThe BAC map is used for sequencing single BACs or BAC-pools covering defined genomic entities by second generation sequencing technologies. Together with the WGS dataset it initiates a sea bass genome sequencing project. This will allow the quantification of polymorphisms through resequencing, which is important for selecting highly performing domesticated fish

Free androgens and progestins and their conjugated forms in serum and urine of stellate sturgeon (Acipenser stellatus Pallas) males

Stellate sturgeon (Acipenser stellatus Pallas) males from Volga River treated with LH-RH-A showed an increase of free, sulfated and glucuronided sex steroid levels in serum and urine at spermiation. Conjugated forms of sex steroids could act as pheromones in sturgeon

Olfactory discrimination of female reproductive status by male tilapia (Oreochromis mossambicus)

The current study investigated whether discrimination of sexual status of female tilapia by males is mediated by olfaction. Size-matched groups of female tilapia were assigned as pre- or post-ovulatory according to the time since their last ovulation (15–19·days pre-ovulatory, N=7; 1–3·days post-ovulatory, N=8). Female-conditioned water and body fluids (urine, bile, faeces and plasma) were assessed for olfactory potency in males by recording the electro-olfactogram (EOG). Water extracts, urine and faeces from pre-ovulatory females all evoked significantly larger amplitude EOGs in male fish (N=6), with correspondingly lower thresholds of detection, than those from post-ovulatory females. Plasma and bile evoked very large amplitude EOGs in males but with no differences between the two groups of females. Anosmic males (N=6) did not behave differently towards pre- or post-ovulatory females, while sham-operated males (N=6) showed a marked increase in urination rate towards pre-ovulatory females. We conclude that the ability of male tilapia to discriminate between females of differing reproductive status is mediated by odorants released into the water, probably via the urine and faeces, by pre-ovulatory females

Levels of free and conjugated androgens and progestins in coelomic fluid and serum of stellate sturgeon (Acipenser stellatus Pallas) females

Stellate sturgeon (Acipenser stellatus Pallas) females from the Volga River were analysed for sex steroids after hormonal stimulation. The results show a significant increase of sulfated sex steroid levels in serum and coelomic fluid (CF) at final maturation (FM). It is hypothesized that the steroid metabolites could be part of a chemical communication system related to reproduction

Olfactory sensitivity to changes in environmental Ca2 in the freshwater teleost Carassius auratus: an olfactory role for the Ca2+ -sensing receptor?

Olfactory sensitivity to changes in environmental Ca2+ has been demonstrated in two teleost species; a salmonid (Oncorhynchus nerka) and a marine/estuarine perciform (Sparus aurata). To assess whether this phenomenon is restricted to species that normally experience large fluctuations in external ion concentrations (e.g. moving from sea water to fresh water) or is present in a much wider range of species, we investigated olfactory Ca2+ sensitivity in the goldfish (Carassius auratus), which is a stenohaline, non-migratory freshwater cyprinid. Extracellular recording from the olfactory bulb in vivo by electroencephalogram (EEG) demonstrated that the olfactory system is acutely sensitive to changes in external Ca2+ within the range that this species is likely to encounter in the wild (0.05–3 mmol l–1). The olfactory system responded to increases in external calcium with increasing bulbar activity in a manner that fitted a conventional Hill plot with an apparent EC50 of 0.9±0.3 mmol l–1 (close to both ambient and plasma free [Ca2+]) and an apparent Hill coefficient of 1.1±0.3 (means ± S.E.M., N=6). Thresholds of detection were below 50 mmol l–1. Some olfactory sensitivity to changes in external [Na+] was also recorded, but with a much higher threshold of detection (3.7 mmol l–1). The olfactory system of goldfish was much less sensitive to changes in [Mg2+] and [K+]. Preliminary data suggest that Ca2+ and Mg2+ are detected by the same mechanism, although with a much higher affinity for Ca2+. Olfactory sensitivity to Na+ may warn freshwater fish that they are reaching the limit of their osmotic tolerance when in an estuarine environment. Olfaction of serine, a potent odorant in fish, was not dependent on the presence of external Ca2+ or Na+. Finally, the teleost Ca2+-sensing receptor (Ca-SR) was shown to be highly expressed in a subpopulation of olfactory receptor neurones by both immunocytochemistry and in situ hybridisation. The olfactory sensitivity to Ca2+ (and Mg2+) is therefore likely to be mediated by the Ca-SR. We suggest that olfactory Ca2+ sensitivity is a widespread phenomenon in teleosts and may have an input into the physiological mechanisms regulating internal calcium homeostasis