The transcriptomic response to copper exposure in the digestive gland of Japanese scallops (Mizuhopecten yessoensis)

doi:10.1016/j.fsi.2015.05.022

Fish & Shellfish Immunology

Volume 46, Issue 2, October 2015, Pages 161-167

https://doi.org/10.1016/j.fsi.2015.05.022 Get rights and content

Highlights

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Gene expression in DG after Cu exposure was illustrated using Illumina platform.
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Genes related to chemical stimulus were detected in DG after Cu exposure.
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KEGG pathways related to immune system and lipid metabolism were detected.

Abstract

The present study was conducted to elucidate the effects of copper exposure on the immune system and lipid metabolism of the Japanese scallop, Mizuhopecten yessoensis. Transcriptional levels of differentially expressed genes (DEGs)in M. yessoensis digestive gland tissue were analyzed using the deep-sequencing platform Illumina HiSeq™ 2000. In total, 841 and 877 genes were identified as significantly up- or down-regulated, respectively. In addition, significant enrichment analysis identified 3 gene ontology terms and 15 pathways involved in the response to copper exposure. Analysis of transcripts related to the immune response revealed a complex pattern of innate recognition receptors, including toll-like receptors, NOD-like receptors and downstream pathway effectors, including those involved in apoptosis. Furthermore, genomic analysis revealed that genes involved in extracellular matrix (ECM)-receptor interactions were enriched in Cu-exposed scallop glands. These results will provide a resource for subsequent gene expression studies regarding heavy metal exposure and the identification of copper-sensitive biomarkers for the aquaculture of M. yessoensis.

Introduction

Heavy metal by products resulting from industrial processes such as mining, smelting, municipal waste removal, petrochemical production, printing manufacturing, and agriculture have been deposited along coastal areas since the Industrial Revolution, and have a significant negative impact on marine ecosystems [1], [2], [3]. Heavy metal pollution has a toxic effect on the surrounding environment, and results in metallic bioaccumulation by marine organisms, particularly in benthic animals [4], [5]. Copper (Cu) is one of the major metal pollutants found in coastal environments. At a low concentration, Cu acts as an essential element and functions in multiple biological processes. It is a cofactor for many enzymes such as cytochrome c oxidase, superoxide dismutase (SOD), dopamine β-hydroxylase and lysyl oxidase, and is involved in electron transport [6]. However, at higher levels, copper is potentially one of the most harmful pollutants, and can be found in elevated concentrations (up to 560 μg/L) from the use of copper sulfate (CuSO₄) in treating macrophyte infestation in aquaculture [7].

The Japanese scallop (Mizuhopecten yessoensis) is widely distributed on the cold coasts of northern China, and has become an attractive species for aquaculture since 1982 due to its relatively large size and high market value [8]. However, the aquaculture of M. yessoensis has been devastated by large-scale death, which has resulted in serious economic losses. Environmental stress, such as excess Cu pollution in coastal waters, may be one of the main factors contributing to the high mortality rates of M. yessoensis [9]. Among the well characterized downstream effects of Cu exposure are the upregulation of reactive oxygen species (ROS) [10], [11], DNA cleavage [12], hepatic lipid metabolism [13], [14], and the induction of apoptotic factors [15]. Most of these studies have been conducted in species of bony fish, however, few reports examine the mechanism of Cu toxicity on bivalves such as M. yessoensis.

One strategy to reduce the high mortality rates of M. yessoensis is to identify disease or stress-resistance genes in scallops, and to use them to genetically alter cultured stocks [16]. Molecular biology techniques, such as expressed sequence tag (EST), serial analysis of gene expression (SAGE), massively parallel signature sequencing (MPSS) and microarray analysis, have been widely used to identify these relevant genes and pathways [16], [17], [18]. Nevertheless, these approaches have several inherent limitations, such as cloning biases, high background levels owing to cross-hybridization and high cost [19], [20]. With the advent of next generation sequencing technologies (NGS), a more comprehensive and accurate transcriptome analysis has become feasible and affordable.

In this study, RNA-Seq (Quantification) [9], a version of NGS, was used for transcriptome quantification and gene expression analysis to determine the effects of Cu exposure to adult of M. yessoensis. The digestive gland was chosen as a target organ for investigation due to the key role it plays in metal accumulation, lipid metabolism, detoxification and immunological function [5], [17], [21]. This study aims to identify genes expressed in the digestive gland that are sensitive to Cu exposure, and to elucidate pathways that can be used to examine the molecular and genetic basis of immunity and stress resistance.

Section snippets

Scallop acquisition

Adult Japanese scallops (M. yessoensis) of both sexes were obtained from Dalian Zhangzidao fishery group (Dalian, Liaoning Province, China). At the beginning of the trial, 30 uniformly sized scallops (shell height: 9.390 ± 0.273 cm, mean ± SD) were stocked in each fiberglass tank for two weeks acclimatization. To minimize the effects of extraneous environmental factors other than Cu exposure, the scallops were cultured in running aerated seawater (salinity 30‰) at 16 ± 1 °C and subjected to

Gene sequencing and mapping

To characterize the response of M. yessoensis to Cu exposure, we performed a high-throughput analysis using RNA-seq (Quantification). We sequenced two cDNA libraries: the control digestive gland group (CD) and the treatment digestive gland group (TD). After removing the dirty reads, the number of clean reads ranged from 4.68 to 4.94 million. The data were deposited in the SRA database (accession number:SRP041735). Reads mapped to a unique sequence are the most critical, as they can explicitly

Acknowledgments

Financial support for this study was provided by Natural Science Foundation of China (Grant no. 31372545), Program for Innovative Research Team (in Science and Technology) in University of Henan Province (Grant no. 14IRTSTHN013), the Science and Technology Key Project of He'nan Educational Committee (Grant no. 14A240001), and by the Ph.D. Foundation of Henan Normal University (Grant no. qd13057).

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Full length articleThe transcriptomic response to copper exposure in the digestive gland of Japanese scallops (Mizuhopecten yessoensis)☆

Highlights

Abstract

Introduction

Section snippets

Scallop acquisition

Gene sequencing and mapping

Acknowledgments

Mar. Pollut. Bull.

Environ. Pollut.

Gene

Comp. Biochem. Phys. C

Am. J. Clin. Nutr.

Aquacult. Eng.

Fish Shellfish Immunol.

Aquat. Toxicol.

Mar. Environ. Res.

Food Chem. Toxicol.

Arch. Environ. Contam. Toxicol.

Aquat. Toxicol.

Aquat. Toxicol.

Methods

Fish Shellfish Immunol.

Biochem. Biophys. Res. Commun.

Aquat. Toxicol.

Aquat. Toxicol.

Dev. Comp. Immunol.

Cancer Lett.

Cytokine Growth Factor Rev.

Ecotoxicol. Environ. Saf.

Immunity

Biochem. Biophys. Res. Commun.

Aquat. Toxicol.

Full length article
The transcriptomic response to copper exposure in the digestive gland of Japanese scallops (Mizuhopecten yessoensis)☆