Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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RNA-seq of polar cod (Boreogadus saida) liver slice culture exposed to benzo[a]pyrene and ethynylestradiol

ABSTRACT: Polar cod, a key fish species in the arctic marine foodweb is vulnerable to effects of pollution from offshore petroleum related activities in the Arctic and sub-arctic region. The study was conducted to map transcriptome responses to in Polar cod (Boreogadus saida) liver slice culture exposed to benzo[a]pyrene (BaP) in the presence or absence of physiological levels of ethynylestradiol (EE2). BaP is a polycyclic aromatic hydrocarbon (PAH), also found in crude oil contaminants. PAHs such as BaP are among the most toxic compounds of crude oil. Precision-cut liver slice cultures from five female polar cod (n = 5/ group, paired design) were exposed to BaP alone (10 µM), or in combination with low concentrations of EE2 (5 nM), to mimic physiological estradiol levels in early vitellogenic female fish. Transcriptome analysis (RNA-seq) was performed after 72 h exposure in culture. The results provide a global view of transcriptome responses to BaP, EE2 and their mixture. In the mixture exposure, BaP resulted attenuation of EE2 stimulated gene expression (anti-estrogenic effects). The results from this ex vivo experiment suggest that pollutants that activate the Ahr pathway such as the PAH compound BaP can result in anti-estrogenic effects that may lead to endocrine disruption in polar cod.

INSTRUMENT(S): TruSeq RNA Sample Preparation protocol, Illumina NovaSeq 6000

ORGANISM(S): Boreogadus saida

SUBMITTER:

PROVIDER: E-MTAB-10509 | biostudies-arrayexpress |

SECONDARY ACCESSION(S): ERP129296

REPOSITORIES: biostudies-arrayexpress

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Project description:Purpose: The objective of this study is to use Atlantic cod (Gadus morhua) PCLS culture and RNA-seq analysis to map gene expression responses to BaP and EE2 exposure in Atlantic cod. Methods: A total of 8 (3 females and 5 males) juvenile cod (G. morhua) (average body weight 498 g) were used for PCLS (n = 6-8 per group). PCLS preparation and chemical exposure in the culture was performed as previously described (Eide et al., 2014). Cod liver slices were cultured in 24-well plates in the growth medium containing DMSO vehicle (CAS No: 67-68-5), BaP (CAS No: 50-32-8) (10 or 1000 nm), EE2 (CAS No: 57-63-6) (10 or 1000 nm) or BaP and EE2 mixture (10 or 1000 nm each). After 48 hors of culture, the slices were collected and snap frozen in liquid nitrogen and stored at -80 °C for RNA extraction. Results: In total, we had 47 high-quality samples, which were aligned to the published CDS of Atlantic cod (ftp://ftp.ensembl.org/pub/release-90/fasta/gadus_morhua/cds/Gadus_morhua.gad) using HISAT2 v2.1.0 (Kim et al., 2015). Counts were generated from the alignments using SAMtools v1.4.1 (Li et al., 2009). Differential expression analysis was performed using edgeR v3.18.1 (McCarthy et al., 2012) between control group and each treated group using paired test, after removing the genes with all-zero counts across all samples of the two compared groups. Differentially expressed genes were defined by p-value < 0.05 after adjustment using the Benjamini-Hochberg multiple testing correction. Several genes were differentially expressed in the BaP and EE2 and mixture treatment groups compared to the group. Hierarchical clustering showed that signature expression profile of top differentially expressed genes in response to the single treatment was distinctly maintained also in the equimolar binary mixtures. Conclusions: Combining high-throughput RNA-seq analysis and PCLS culture in Atlantic cod, we have shown that BaP and EE2 differentially regulated many genes in the AHR and ER pathways. The results show that omics data can be generated using an efficient PCLS in vitro culture system.

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RNA-seq of polar cod (Boreogadus saida) liver slice culture exposed to benzo[a]pyrene and ethynylestradiol

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