Differentially expressed lncRNAs involved in immune responses of Haliotis diversicolor and H. discus hannai challenged with Vibrio parahaemolyticus

https://doi.org/10.1016/j.cbd.2021.100873Get rights and content

Highlights

  • This is the first study that identified lncRNAs related to V. parahaemolyticus infection in abalone.

  • Some immune-related genes (including NFKB1, NFKB2, lectin C) and lncRNAs were validated by real-time PCR.

  • The unique and common lncRNA and mRNA in two abalone species were identified for the first time.

Abstract

Although many studies have shown that lncRNA, a non-coding RNA with a length of more than 200 bases, is involved in various biological functions, including the immune process, stress process, and cell development process. However, the function of lncRNA in abalone, especially in immunity, has been rarely studied. H. discus hannai and H. diversicolor are two main aquaculture abalone, and their growth is easily affected by the main pathogen Vibrio parahaemolyticus. Through rigorous screening procedures for transcripts in this study, we found that lncRNAs were 34,240, 23,022 in Haliotis diversicolor and H. discus hannai injected with V. parahaemolyticus, respectively. We also identified the unique and common lncRNAs and mRNAs of two abalone species for the first time; the shared lncRNAs and mRNAs in Haliotis diversicolor and H. discus hannai were 2352 and 13,165, respectively. Then gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of the differentially expressed target genes of common and unique lncRNAs has shown that common lncRNAs could be widely involved in the biological processes of stress and cell development in both abalone species. In contrast, unique lncRNAs are linked to the Toll-like receptor, NF-kappaB signaling pathway of H. diversicolor, and pattern recognition receptors and lectins immune-related pathways of H. discus hannai. The co-expression network shows that some immune-related genes, such as INFK1, INFK2, CASP2, CASP8, IRAK1, lectin C, were closely related to lncRNAs. Further, we identified the targeted relationship between some immune-related genes and lncRNAs by qRT-PCR, through which we showed that the expression trend between targeted genes, such as INFK1 and Lnc7057, lectin C and Lnc6943, Lnc5637, and PLCG1 and Lnc1692, were consistent. In general, our results showed that lncRNA expression was induced in the two species of abalone after being infected with V. parahaemolyticus, and lncRNA was involved in the immune response of abalone by targeting coding genes.

Introduction

LncRNA is a non-coding RNA with a length of more than 200 bases(Isin et al., 2014). lncRNA often originated from the intron or exon promoter region (Valadkhan and Gunawardane, 2016) of some coding genes through gene locating. According to different characteristics of lncRNAs, lncRNAs can be classified into cis-lncRNAs, and trans-lncRNAs (Ma et al., 2013). cis-LncRNAs can regulate the same allele's target genes, which are close to lncRNAs (Dykes and Emanueli, 2017). trans-LncRNAs can guide the changes of coding genes far away and drive the chromatin changes of the nucleus by binding to target genes in the way of RNA heteroduplex or RNA:DNA:DNA triplex (Wang and Chang, 2011). At the same time, lncRNAs may be involved in the splicing, translation, and degradation in the process of post-transcription (Shi et al., 2015).

LncRNAs can regulate many biological processes such as stress response (Huo et al., 2020; Li et al., 2018), cell development (Huang et al., 2018), Oxidation-Reduction reactions (Huang et al., 2019) in different aquatic animals. In addition, lncRNAs play an important role in the immune response of large yellow croaker, Epinephelus coioides, Penaeus vannamei, and sea cucumber (Mu et al., 2016; Liu et al., 2019; Tang et al., 2019; Ren et al., 2020). Although studies have shown that lncRNAs were extensively involved in the immune response of other aquatic animals, while studies on the immune regulation of non-model organisms, especially marine mollusks, are still relatively rare.

Haliotis diversicolor and H. discus hannai are two different most important aquaculture abalone species in China and have high commercial value. However, their aquaculture industry suffers from different diseases cuasing mass mortality of cultured abalone due to the increasing density and scale of mariculture. V. parahaemolyticus is a prominent pathogenic microorganism in marine, estuarine and aquacultural ecosystems (Srinivasan et al., 2020), and it is one of the main pathogenic bacteria causing vibriosis that results in high morbidity and mortality in aquaculture (Liu and De Mitcheson, 2008). The outbreak of pathogenic infectious diseases dominated by V. parahaemolyticus would cause a large number of abalone deaths (Cheng et al., 2004; Hasrimi et al., 2018). Therefore, the purpose of our study is to explore the role of lncRNA in the immunomodulatory response of abalone injected with V. parahaemolyticus.

Our study was the first to identify the lncRNA expression profiles of H. diversicolor and H. discus hannai infected by V. parahaemolyticus. By screening the unique and common lncRNAs in two abalone species and exploring their function, it was found that lncRNAs were widely involved in the immune process, cell differentiation process, and stress process of the two species of abalones by targeting mRNAs. Meanwhile, we also identified some lncRNAs and their target genes that may be related to the immune response in the two species of abalone. Our results may provide essential resources for further understanding the interaction mechanism between lncRNAs-mRNAs in abalone and help explore the molecular function of lncRNAs in the immune response of abalone infected by V. parahaemolytic.

Section snippets

Animals and sample preparation

Two different adult abalone with a weight of 16.45 ± 2.50 g and body length of 6.00 ± 0.50 cm were obtained from the Peiyang abalone plant (Xiamen, Fujian Province). As described in our previous publication (Sun et al., 2016), these abalones were remained at a flowing air stream and a circulating seawater system (salinity was kept at 30‰) with a constant temperature of 25 °C and dissolved oxygen (DO) of 6.2 mg/L, and they were fed with Laminaria japonica once a day. Seawater temperature and DO

Sequence assembly and identification of lncRNA characterization

After sequencing and bioinformatics analysis of the hemocyte samples, 158,937,667 and 170,082,258 raw reads were generated in H. discus hanai and H. diversicolor, respectively. After removing adaptor and low-quality reads, 154,241,843 (97.04%) and 165,613,084 (97.37%) clean reads for the two abalone species were obtained, respectively. Then the assembly of these clean reads was performed, and 367,162 contigs were produced in the H. diversicolor transcriptome, and 316,734 contigs were produced

Discussion

LncRNA as a non-coding RNA molecule has been found to play an important role in various biological processes, and its role in immunity has become one of the focuses of people studies. Of note, Valenzuela-Muñoz et al. (2019) reported that lncRNAs were involved in the immune defense process of zebrafish infected with Spring viremia of carp virus via targeting adjacent coding genes. Sun and Feng (2018) also showed that lncRNA was widely involved in the initial antiviral immune response of

Conclusion

We identified the expression profile of lncRNAs for the first time in H. diversicolor and H. discus hannai infected with V. parahaemolyticus. It is also the first time use of the non-referenced transcriptome method for comparing the functional difference of lncRNAs in H. diversicolor and H. discus hannai. This study found some differentially expressed lncRNAs related to the genes involved in immune response in abalone infected with V. parahaemolyticus. The result indicated that lncRNAs might

Declaration of competing interest

All authors have read and approved the manuscript. None of the authors had a conflict of interest that interfered with our work.

Acknowledgments

This research was funded by the National Key Research and Development Program of China (Grant Number: 2018YFD0900304–5, 2021YFE0106100); Discipline Development Grant from College of Animal Sciences FAFU (712018R0404), Open fund project of Fujian Engineering Research Center of Aquatic Breeding and Healthy Aquaculture (DF20902); the Natural Science Foundation of China (No. 41176152); International Science and Technology Cooperation and Communication Grant of Fujian Agriculture and Forestry

CRediT authorship contribution statement

G.L participated in the experimental design, experimental data processing and drawing, and wrote the manuscript. C·Y participated in the experimental data processing and qRT-PCR drawing, X.Z and Y·S designed the experiment and participated in the sample collection, Z.Z and Y·W edited the manuscript and supervised the study. All the authors reviewed the manuscript.

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