Alternative splicing is highly variable among Daphnia pulex lineages in response to acute copper exposure

Sneha Suresh,Frédéric J J Chain,Teresa J Crease,Melania E Cristescu

doi:10.1186/s12864-020-06831-4

Sneha Suresh, Frédéric J J Chain + Show 2 more

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https://doi.org/10.1186/s12864-020-06831-4

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Abstract

BackgroundDespite being one of the primary mechanisms of gene expression regulation in eukaryotes, alternative splicing is often overlooked in ecotoxicogenomic studies. The process of alternative splicing facilitates the production of multiple mRNA isoforms from a single gene thereby greatly increasing the diversity of the transcriptome and proteome. This process can be important in enabling the organism to cope with stressful conditions. Accurate identification of splice sites using RNA sequencing requires alignment to independent exonic positions within the genome, presenting bioinformatic challenges, particularly when using short read data. Although technological advances allow for the detection of splicing patterns on a genome-wide scale, very little is known about the extent of intraspecies variation in splicing patterns, particularly in response to environmental stressors. In this study, we used RNA-sequencing to study the molecular responses to acute copper exposure in three lineages of Daphnia pulex by focusing on the contribution of alternative splicing in addition to gene expression responses.ResultsBy comparing the overall gene expression and splicing patterns among all 15 copper-exposed samples and 6 controls, we identified 588 differentially expressed (DE) genes and 16 differentially spliced (DS) genes. Most of the DS genes (13) were not found to be DE, suggesting unique transcriptional regulation in response to copper that went unnoticed with conventional DE analysis. To understand the influence of genetic background on gene expression and alternative splicing responses to Cu, each of the three lineages was analyzed separately. In contrast to the overall analysis, each lineage had a higher proportion of unique DS genes than DE genes suggesting that genetic background has a larger influence on DS than on DE. Gene Ontology analysis revealed that some pathways involved in stress response were jointly regulated by DS and DE genes while others were regulated by only transcription or only splicing.ConclusionsOur findings suggest an important role for alternative splicing in shaping transcriptome diversity in response to metal exposure in Daphnia, highlighting the importance of integrating splicing analyses with gene expression surveys to characterize molecular pathways in evolutionary and environmental studies.

Highlights

Despite being one of the primary mechanisms of gene expression regulation in eukaryotes, alternative splicing is often overlooked in ecotoxicogenomic studies
A total of 588 differentially expressed (DE) genes were identified between all 15 samples exposed to acute Cu stress versus all 6 control samples (FDR corrected p-value < 0.05)
Of the DE genes, 20 genes had an False Discovery Rate (FDR) corrected p-value < 0.01 and an absolute log2 fold change > 4, which we identified as having the most extreme expression differences between copper and controls (Fig. 1b)

Summary

Introduction

Despite being one of the primary mechanisms of gene expression regulation in eukaryotes, alternative splicing is often overlooked in ecotoxicogenomic studies. The process of alternative splicing facilitates the production of multiple mRNA isoforms from a single gene thereby greatly increasing the diversity of the transcriptome and proteome This process can be important in enabling the organism to cope with stressful conditions. We used RNA-sequencing to study the molecular responses to acute copper exposure in three lineages of Daphnia pulex by focusing on the contribution of alternative splicing in addition to gene expression responses. Anthropogenic activities such as mining and intensive agriculture have led to a substantial amount of heavy metal pollution in aquatic ecosystems [1]. Recent genomewide studies have provided important insights into the molecular basis of transcription in response to metals, but much less is known about the contribution of other mechanisms that regulate expression via RNA processing such as alternative splicing

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