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Identification of differentially expressed Oncorhynchus mykiss microRNA in plasma, mucus, and water following acute stress using high throughput sequencing.

ABSTRACT: Circulating plasma microRNAs (miRNAs) are well established as biomarkers of several diseases in humans and have recently been used as indicators of environmental exposures in fish. However, the role of plasma miRNAs in regulating acute stress responses in fish is largely unknown. Tissue and plasma miRNAs have recently been associated with excreted miRNAs in humans however external miRNAs have never been measured in fish. The objective of this study was to characterize the plasma miRNA profile in response to acute stress in rainbow trout (Oncorhynchus mykiss), as well as miRNA profiles in novel external samples, (fish epidermal mucus and the surrounding water). RNA was extracted and sequenced from plasma, mucus, and water collected from rainbow trout and their surrounding environment prior to and one-hour following a three-minute air exposure, a known inducer of an acute stress response in fish. Following small RNA-Seq and pathway analysis, we identified differentially expressed plasma miRNAs that targeted biosynthetic, degradation, and metabolic pathways. We successfully isolated miRNA from trout mucus and the surrounding water and detected differences in miRNA expression one-hour post air stress. The altered miRNA profiles in mucus and water were unique to the altered plasma miRNA profile, indicating that the plasma miRNA response was not associated with or immediately reflected in external samples. This research expands our understanding of the role of plasma miRNA in the acute stress response of fish and is the first study to report on the successful isolation and profiling of miRNA from fish mucus and water samples. Measurements of miRNA from plasma, mucus, and water can be further studied and have the potential to be applied in environmental monitoring as non-lethal indicators of acute stress in fish.

ORGANISM(S): Oncorhynchus mykiss

PROVIDER: GSE151138 | GEO | 2020/12/28

REPOSITORIES: GEO

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Project description:Hypoxia negatively affects the behavior, growth, reproduction, and survival of fish, causing serious economic losses to aquaculture. Rainbow trout (Oncorhynchus mykiss), an important economic fish worldwide, belongs to a hypoxia-sensitive fish species. However, the regulatory mechanisms of miRNAs under hypoxia stress response of rainbow trout remains unclear. In this study, rainbow trout were subjected to hypoxia stress (DO: 3.5 mg/L) for 3 h (H3h_L), 12 h (H12h_L), 24 h (H24h_L) and 3 h reoxygenation (R3h_L) to systemically evaluate the changes of miRNA expression profiles in liver, and the functions of sha-miR-92a_L+2R+4 were investigated. We found 17, 144, 57 and 55 differentially expressed (DE) miRNAs in H3h_L vs. control (N_L), H12h_L vs. N_L, H24h_L vs. N_L and R3h_L vs. N_L comparisons, respectively. Enrichment analysis revealed that the targets of these DE miRNAs were significantly enriched in HIF signaling pathway, VEGF signaling pathway, FoxO signaling pathway and glycolysis/gluconeogenesis. Through miRNA-mRNA nteraction and weighted gene co-expression network analysis (WGCNA), five key DE miRNAs (sha-miR-92a_L+2R+4, ssa-miR-128-3p, ssa-miR-101b-3p_R+1, ola-miR-199a-5p_R+2 and tni-miR-199_1ss18CG) were identified, which can target at least two hypoxia-responsive genes, such as vegfaa, ho, glut1a and junb. Functional analysis found that sha-miR-92a_L+2R+4 directly regulated vegfaa expression by targeting its 3′-UTR, overexpression of sha-miR-92a_L+2R+4 significantly decreased vegfaa expression in rainbow trout liver cells, while opposite results were obtained after transfection of sha-miR-92a_L+2R+4 inhibitor. Furthermore, overexpression of sha-miR-92a_L+2R+4 promoted rainbow trout liver cell proliferation and inhibited apoptosis. These results deepen our understanding of the crucial roles of miRNAs under hypoxia stress in rainbow trout, and provide valuable information for further studying the regulatory mechanisms of key hypoxia-responsive miRNAs and breeding hypoxia-tolerant rainbow trout species.

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Identification of differentially expressed Oncorhynchus mykiss microRNA in plasma, mucus, and water following acute stress using high throughput sequencing.

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