A novel hypoxic lncRNA, LOC110520012 sponges miR-206-y to regulate angiogenesis and liver cell proliferation in rainbow trout (Oncorhynchus mykiss) by targeting vegfaa.

Long non-coding RNA (lncRNA) is a novel emerging type of competitive endogenous RNA (ceRNA) that performs key functions in multiple biological processes. However, little is known about the roles of lncRNA under hypoxia stress in fish. Here, vascular endothelial growth factor-Aa (vegfaa) was cloned in rainbow trout (Oncorhynchus mykiss), with the complete cDNA sequence of 2914 bp, encoding 218 amino acids. The molecular weight of the protein was approximately 25.33 kDa, and contained PDGF and VEGF_C domains. Time-course and spatial expression patterns revealed that LOC110520012 was a key regulator of rainbow trout in response to hypoxia stress, and LOC110520012, miR-206-y and vegfaa exhibited a ceRNA regulatory relationship in liver, gill, muscle and rainbow trout liver cells treated with acute hypoxia. Subsequently, the targeting relationship of LOC110520012 and vegfaa with miR-206-y was confirmed by dual-luciferase reporter analysis, and overexpression of LOC110520012 mediated the inhibition of miR-206-y expression in rainbow trout liver cells, while the opposite results were obtained after LOC110520012 silencing with siRNA. We also proved that vegfaa was a target of miR-206-y in vitro and in vivo, and the vegfaa expression and anti-proliferative effect on rainbow trout liver cells regulated by miR-206-y mimics could be reversed by LOC110520012. These results suggested that LOC110520012 can positively regulate vegfaa expression by sponging miR-206-y under hypoxia stress in rainbow trout, which facilitate in-depth understanding of the molecular mechanisms of fish adaptation and tolerance to hypoxia.

Dietary supplementation of Chinese herbal medicines enhances the immune response and resistance of rainbow trout (Oncorhynchus mykiss) to infectious hematopoietic necrosis virus.

Rainbow trout is a widely farmed economical cold-water fish worldwide, but the prevalence of infectious hematopoietic necrosis virus (IHNV) presents a severe risk to the aquaculture industry, resulting in high mortality and huge economic losses. In this study, the impacts of different concentrations (0, 10, 20, and 30 g/kg) of Chinese herbal medicine mixture (CHMM) on the immune response and resistance of rainbow trout to IHNV infection were evaluated. The results show that CHMM noticeably increased (P < 0.05) T-SOD, CAT, AST, ALT, ACP, and AKP activities and decreased MDA content. NF-κB, TNF-α, IFN-ß, IL-1ß, JAK1, HSP70, and HSP90 expressions were significantly upregulated (P < 0.05) in all CHMMs, while SOCS2 expression was downregulated (P < 0.05). Following infection with IHNV, feeding rainbow trout with varying amounts of CHMM resulted in noticeably increased (P < 0.05) T-SOD, ACP, and AKP activities and significantly decreased (P < 0.05) MDA content and AST and ALT activities. TNF-α, IFN-ß, IL-1ß, HSP70, and HSP90 expressions were significantly upregulated (P < 0.05) in all CHMMs, while the expressions of JAK1 and SOCS2 were downregulated. The expression level of the IHNV G protein gene at a dosage of 20 g/kg was notably lower than that of the other CHMM feeding groups. This study provides a solid scientific basis for promoting CHMM as an immunostimulant for boosting antiviral immunity in rainbow trout.

Comparative transcriptomics combined with physiological and functional analysis reveals the regulatory mechanism of rainbow trout (Oncorhynchus mykiss) under acute hypoxia stress.

Hypoxia, largely triggered by global warming and water contamination, has become an environmental issue of great concern, posing a great threat to aquatic ecosystem. As one of the world's most economically important fish, rainbow trout (Oncorhynchus mykiss) is extremely intolerant of hypoxic environments, however, little is known about the roles of non-coding RNAs (ncRNAs) in the response of rainbow trout to hypoxia stress. Herein, effects of moderate (Tm12L) and severe hypoxia for 12 h (Ts12L) and 12 h reoxygenation on histology, biochemical parameters (antioxidant, metabolism and immunity) and transcriptome (lncRNA, miRNA and mRNA) in rainbow trout liver were investigated. We further validated the regulatory relationships between LOC110519952, novel-m0023-5p and glut1a via dual­luciferase reporter, overexpression and silencing assays. Compared with Tm12L, the liver in Ts12L showed more severe oxidative damage. Anaerobic, lipid and protein metabolism was enhanced under hypoxia stress, especially in Ts12L. We also found that Tm12L could strengthen innate immune response, which was inhibited in Ts12L. Besides, several hypoxia-related genes (glut1a, vegfaa, hmox, epoa, foxo1a and igfbp1) and ceRNA networks were identified from 1824, 427 and 545 differentially expressed mRNAs, miRNAs and lncRNAs, including LOC118965299-novel-m0179-3p-epoa, LOC110519952-novel-m0023-5p-glut1a, MSTRG.7382.2-miR-184-y-hmox and LOC110520012-miR-206-y-vegfaa. Through in vitro and in vivo functional analysis, we demonstrated that glut1a is a target of novel-m0023-5p, and LOC110519952 can positively regulate glut1a by targeting novel-m0023-5p. Introduction of LOC110519952 could attenuate the promoting effects of novel-m0023-5p on rainbow trout liver cell viability and proliferation. This study highlights the differences in the regulatory mechanism of rainbow trout under different concentrations of hypoxia stress and provides valuable data for further research on the molecular mechanisms of fish adaptation to hypoxic environments.

Antiviral effect of miR-206 in rainbow trout (Oncorhynchus mykiss) against infectious hematopoietic necrosis virus infection.

Infectious hematopoietic necrosis (IHN), caused by IHN virus, is a highly contagious and lethal disease that seriously hampers the development of rainbow trout (Oncorhynchus mykiss) aquaculture. However, the immune response mechanism of rainbow trout underlying IHNV infection remains largely unknown. MicroRNAs act as post-transcriptional regulators of gene expression and perform a crucial role in fish immune response. Herein, the regulatory mechanism and function of miR-206 in rainbow trout resistance to IHNV were investigated by overexpression and silencing. The expression analysis showed that miR-206 and its potential target receptor-interacting serine/threonine-protein kinase 2 (RIP2) exhibited significant time-dependent changes in headkidney, spleen and rainbow trout primary liver cells infected with IHNV and their expression displayed a negative correlation. In vitro, the interaction between miR-206 and RIP2 was verified by luciferase reporter assay, and miR-206 silencing in rainbow trout primary liver cells markedly increased RIP2 and interferon (IFN) expression but significantly decreased IHNV copies, and opposite results were obtained after miR-206 overexpression or RIP2 knockdown. In vivo, overexpressed miR-206 with agomiR resulted in a decrease in the expression of RIP2 and IFN in liver, headkidney and spleen. This study revealed the key role of miR-206 in anti-IHNV, which provided potential for anti-viral drug screening in rainbow trout.

Involvement of miR-495 in the skin pigmentation of rainbow trout (Oncorhynchus mykiss) through the regulation of mc1r.

MicroRNAs (miRNAs) play crucial roles in skin pigmentation in animals. Rainbow trout (Oncorhynchus mykiss) is a key economic fish species worldwide, and skin color directly affects its economic value. However, the functions of miRNAs in rainbow trout skin pigmentation remain largely unknown. Herein, we overexpressed and silenced miR-495 in vitro and in vivo to investigate its functions. The analysis of spatial and temporal expression patterns suggested that miR-495 is a potential regulator during the process of skin pigmentation. In vitro, mc1r was validated as a direct target for miR-495 by dual-luciferase reporter assay, and overexpression of miR-495 significantly inhibited mc1r expression; in contrast, mc1r and its downstream gene mitf levels were markedly upregulated by decreased miR-495. In vivo, overexpressed miR-495 by injecting agomiR-495 led to a substantial decrease in the expression of mc1r and mitf in dorsal skin and liver, while the opposite results were obtained after miR-495 silencing by antagomiR-495. These findings suggested that miR-495 can target mc1r to regulate rainbow trout skin pigmentation, which provide a potential basis for using miRNAs as target drugs to treat pigmentation disorders and melanoma.

Integrative mRNA-miRNA interaction analysis associated with the immune response in the head kidney of rainbow trout (Oncorhynchus mykiss) after infectious hematopoietic necrosis virus infection.

Rainbow trout (Oncorhynchus mykiss) is an important cold-water fish widely cultivated in China. The frequent occurrence of viral diseases caused by infectious hematopoietic necrosis virus (IHNV) seriously restricted the healthy development of the rainbow trout farming industry. However, the immune defense mechanism induced by IHNV in rainbow trout has not been fully elucidated. In the present study, we detected mRNA and miRNA expression profiles in rainbow trout head kidney after IHNV infection using RNA-seq and identified key immune-related genes and miRNAs. The results showed that a total of 7486 genes and 277 miRNAs were differentially expressed, and numerous differentially expressed genes (DEGs) enriched in the immune-related pathways such as Toll-like receptor signaling pathway, RIG-I-like receptor signaling pathway, NOD-like receptor signaling pathway, cytokine-cytokine receptor interaction, and JAK-STAT signaling pathway were significantly up-regulated, including LGP2, MDA5, TRIM25, IRF3, IRF7, TLR3, TLR7, TLR8, MYD88, and IFN1. Integration analysis identified six miRNAs (miR-141-y, miR-200-y, miR-144-y, miR-2188-y, miR-725-y, and miR-203-y) that target at least six key immune-related genes (TRIM25, LGP2, TLR3, TLR7, IRF3, and IRF7). Further, we verified selected immune-related mRNAs and miRNAs through qRT-PCR and confirmed the reliability of the RNA-seq results. These findings improve our understanding of the immune mechanism of rainbow trout infected with IHNV and provide basic data for future breeding for disease resistance in rainbow trout.

miR-330 targeting BCO2 is involved in carotenoid metabolism to regulate skin pigmentation in rainbow trout (Oncorhynchus mykiss).

BACKGROUND: MicroRNAs (miRNAs) play a critical role in regulating skin pigmentation. As a key economic trait, skin color directly affects the market value of rainbow trout (Oncorhynchus mykiss), however, the regulatory mechanism of most miRNAs in fish skin color is still unclear. RESULTS: In this study, the full-length cDNA sequence of ß-carotene oxygenase 2 (BCO2, a key regulator of carotenoid metabolism) from the rainbow trout was obtained using rapid-amplification of cDNA ends (RACE) technology, and qRT-PCR was used to investigate the differential expression of miR-330 and BCO2 in 14 developmental stages and 13 tissues between wild-type rainbow trout (WTrt) and yellow mutant rainbow trout (YMrt). Additionally, the function of miR-330 was verified by overexpression and silencing in vitro and in vivo. The results showed that the complete cDNA sequence of BCO2 was 2057 bp with a 1707 bp ORF, encoding a 568 amino acid protein having a molecular weight of 64.07 kD. Sequence alignment revealed that higher conservation of BCO2 protein amongst fishes than amongst other vertebrates, which was further confirmed by phylogenetic analysis. The analysis of spatial and temporal expression patterns suggested that BCO2 and miR-330 were abundantly expressed from fertilized-stage to multi-cell as well as in the dorsal and ventral skin of WTrt and YMrt, and their expression patterns were opposite in most of the same periods and tissues. In vitro, luciferase reporter assay confirmed that BCO2 was a direct target of miR-330, and transfection of miR-330 mimics into rainbow trout liver cells resulted in a decrease in the expression of BCO2; conversely, miR-330 inhibitor had the opposite effect to the miR-330 mimics. In vivo, miR-330 agomir significantly decreased BCO2 expression in dorsal skin, tail fin, and liver. Furthermore, overexpression of miR-330 could suppress cell proliferation and induce apoptosis. CONCLUSION: Our results showed that miR-330 is involved in the regulation of skin pigmentation in rainbow trout by targeting BCO2 and shows its promise as a potential molecular target to assist the selection of rainbow trout with better skin color patterns.

Comprehensive analysis of immune parameters, mRNA and miRNA profiles, and immune genes expression in the gill of rainbow trout infected with infectious hematopoietic necrosis virus (IHNV).

Rainbow trout (Oncorhynchus mykiss) is a species of cold-water fish with important economic values, widely cultivated worldwide. However, the outbreak of infectious hematopoietic necrosis virus (IHNV) caused the large-scale death of rainbow trout and seriously restricted the development of the trout farming industry. In this study, the changes of immune parameters in different periods (6-, 12-, 24-, 48-, 72-, 96-, 120-, and 144 h post-infection (hpi)), transcriptome profiles of 48 hpi (T48G) compared to control (C48G), and key immune-related genes expression patterns were measured in rainbow trout gill following IHNV challenge through biochemical methods, RNA sequencing (RNA-seq), and quantitative real-time polymerase chain reaction (qRT-PCR). The results showed that alkaline phosphatase (AKP), acid phosphatase (ACP), total superoxide dismutase (T-SOD), alanine aminotransferase (ALT), and aspartate aminotransferase (AST) activities, as well as lysozyme (LZM) and malonaldehyde (MDA) content decreased and then increased during infection, and remained at a high level after 48 hpi (P < 0.05), whereas catalase (CAT) activity showed a significant peak at 48 hpi (P < 0.05). The mRNA and miRNA analysis identified 4343 differentially expressed genes (DEGs) and 11 differentially expressed miRNAs (DEMs), and numerous immune-related DEGs involved in the Toll-like receptor signaling pathway, apoptosis, DNA replication, p53 signaling, RIG-I-like receptor signaling pathway, and NOD-like receptor signaling pathway and expression were significantly up-regulated in T48Gm group, including tlr3, tlr7, tlr8, traf3, ifih1, trim25, dhx58, ddh58, hsp90a.1, nlrc3, nlrc5, socs3, irf3, irf7, casp7, mx1, and vig2. The integrated analysis identified several important miRNAs (ola-miR-27d-3p_R+5, gmo-miR-124-3-5p, ssa-miR-301a-5p_L+2, and ssa-miR-146d-3p) that targeted key immune-related DEGs. Expression analysis showed that tlr3, tlr7, traf3, ifih1, dhx58, hap90a.1, irf3, irf7, and mx1 genes increased and then decreased during infection, and peaked at 72 hpi (P < 0.05). However, trim25 expression peaked at 96 hpi (P < 0.05). This study contributes to understanding immune response of rainbow trout against IHNV infection, and provides new insights into the immune regulation mechanisms and disease resistance breeding studies.

Integrative analysis of miRNA and mRNA expression associated with the immune response in the intestine of rainbow trout (Oncorhynchus mykiss) infected with infectious hematopoietic necrosis virus.

Rainbow trout (Oncorhynchus mykiss), an economically important cold-water fish cultured worldwide, suffers from infectious hematopoietic necrosis virus (IHNV) infection, resulting in huge financial losses. In order to understand the immune response of rainbow trout during virus infection, we explored trout intestine transcriptome profiles following IHNV challenge, and identified 3355 differentially expressed genes (DEGs) and 80 differentially expressed miRNAs (DEMs). Transcriptome analysis revealed numerous DEGs involved in immune responses, such as toll-like receptor 3 (TLR3), toll-like receptor 7/8 (TLR7/8), tripartite motif-containing 25 (TRIM25), DExH-Box helicase 58 (DHX58), interferon-induced with helicase C domain 1 (IFIH1), interferon regulatory factor 3 (IRF3/7), signal transducer and activator of transcription 1 (STAT1) and heat shock protein 90-alpha 1 (HSP90A1). Integrated analysis identified five key miRNAs (miR-19-y, miR-181-z, miR-203-y, miR-143-z and miR-206-y) targeting at least two important immune genes (TRIM25, DHX58, STAT1, TLR7/8 and HSP90A1). Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses showed that DEGs and target genes were significantly enriched in various immune-related terms including immune system process, binding, cell part and pathways of Toll-like receptor signalling, RIG-I-like receptor signalling, NOD-like receptor signalling, JAK-STAT signalling, PI3K-Akt signalling, NF-kappa B signalling, IL-17 signalling and AGE-RAGE signalling. In addition, protein-protein interaction networks (PPI) was used to display highly interactive DEG networks involving eight immune-related pathways. The expression trends of 12 DEGs and 10 DEMs were further verified by quantitative real-time PCR, which confirmed the reliability of the transcriptome sequencing results. This study expands our understanding of the immune response of rainbow trout infected with IHNV, and provides valuable resources for future studies on the immune molecular mechanism and disease resistance breeding.

Integrated analysis of immune parameters, miRNA-mRNA interaction, and immune genes expression in the liver of rainbow trout following infectious hematopoietic necrosis virus infection.

Rainbow trout (Oncorhynchus mykiss) is an important economical cold-water fish worldwide. However, infection with infectious hematopoietic necrosis virus (IHNV) has severely restricted the development of aquaculture and caused huge economic losses. Currently, little is known about the immune defense mechanisms of rainbow trout against IHNV. In this study, we detected the changes of immune parameters over different post-infection periods (6-, 12-, 24-, 48-, 72-, 96-, 120-, and 144 hours post-infection (hpi)), mRNA and miRNA expression profiles under 48 hpi (T48L) compared to control (C48L), and key immune-related genes expression patterns in rainbow trout liver following IHNV challenge through biochemical methods, RNA-seq, and qRT-PCR, and the function of miR-330-y was verified by overexpression and silencing in vitro and in vivo. The results revealed that alkaline phosphatase (AKP), alanine aminotransferase (ALT), catalase (CAT), and total superoxide dismutase (T-SOD) activities, and lysozyme (LZM) content showed significant peaks at 48 hpi, whereas malondialdehyde (MDA) content and aspartate aminotransferase (AST) activity decreased continuously during infection, and acid phosphatase (ACP) activity varied slightly. From RNA-seq, a total of 6844 genes and 86 miRNAs were differentially expressed, and numerous immune-related differentially expressed genes (DEGs) involved in RIG-I-like receptor signaling pathway, Toll-like receptor signaling pathway, NOD-like receptor signaling pathway, cytokine-cytokine receptor interaction, and antigen processing and presentation were significantly upregulated in T48Lm group, including IFIH1, DHX58, MAVS, TRAF3, IRF3, IRF7, MX1, TLR3, TLR8, MYD88, NOD1, NOD2, IL-8, CXCR1, CD209, CD83, and TAP1. Integrated analysis identified seven miRNAs (miR-425-x, miR-185-x, miR-338-x, miR-330-y, miR-361-x, miR-505-y, and miR-191-x) that target at least three key immune-related DEGs. Expression analysis showed that IFIH1, DHX58, IRF3, IRF7, MX1, TLR3, TLR8, and MYD88 showed a marked increase after 24 hpi during infection. Further research confirmed TAP1 as one of the targets of miR-330-y, overexpression of miR-330-y with mimics or agomir significantly reduced the expression levels of TAP1, IRF3, and IFN, and the opposite effects were obtained by inhibitor. These results facilitate in-depth understanding of the immune mechanisms in rainbow trout against IHNV.

MiR-382 Functions on the Regulation of Melanogenesis via Targeting dct in Rainbow Trout (Oncorhynchus mykiss).

MicroRNAs (miRNAs) are a class of endogenous small noncoding RNAs that are involved in a diverse collection of biological processes as important post-transcriptional regulators. However, little is known about the molecular regulatory mechanism of miRNAs in fish skin pigmentation. In this study, we first confirmed that dopachrome tautomerase (dct), a key gene of melanogenesis, is a target of miR-382 in rainbow trout (Oncorhynchus mykiss) using luciferase reporter assay. The analysis of different developmental stages and tissue expression patterns between wild-type and yellow mutant rainbow trout suggested that miR-382 is a potential regulator during the process of skin pigmentation. In vitro, miR-382 mimics in rainbow trout primary liver cells significantly downregulated dct expression and resulted in decreased expression of key melanogenic genes including tyrosine-related protein 1 (tyrp1) and premelanosome protein (pmel), whereas the expression level of dct was markedly increased after transfected with miR-382 inhibitor. In vivo, overexpression of miR-382 by injection of miR-382 agomir significantly depressed the expression of dct in dorsal skin, tail fin, and liver and then reduced the expression levels of tyrp1 and pmel. Furthermore, transfection of miR-382 mimics inhibited cell proliferation and induced apoptosis. Taken together, our results identified a functional role of miR-382 in rainbow trout skin pigmentation through targeting dct, which facilitate understanding the regulatory mechanism of rainbow trout skin color at the post-transcriptional level and provide a theoretical basis for molecular breeding with skin color as the target trait.

Integrated Analysis of lncRNA and circRNA Mediated ceRNA Regulatory Networks in Skin Reveals Innate Immunity Differences Between Wild-Type and Yellow Mutant Rainbow Trout (Oncorhynchus mykiss).

Fish skin is a vital immune organ that forms the first protective barrier preventing entry of external pathogens. Rainbow trout is an important aquaculture fish species that is farmed worldwide. However, our knowledge of innate immunity differences between wild-type (WR_S) and yellow mutant rainbow trout (YR_S) remains limited. In this study, we performed whole transcriptome analysis of skin from WR_S and YR_S cultured in a natural flowing water pond. A total of 2448 mRNAs, 1630 lncRNAs, 22 circRNAs and 50 miRNAs were found to be differentially expressed (DE). Among these DEmRNAs, numerous key immune-related genes, including ifih1, dhx58, trim25, atp6v1e1, tap1, tap2, cd209, hsp90a.1, nlrp3, nlrc3, and several other genes associated with metabolism (gstp1, nampt, naprt and cd38) were identified. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses of DEmRNAs revealed that many were significantly enriched in innate immune-related GO terms and pathways, including NAD+ADP-ribosyltransferase activity, complement binding, immune response and response to bacterium GO terms, and RIG-I-like receptor signaling, NOD-like receptor signaling and phagosome KEGG pathways. Furthermore, the immune-related competing endogenous RNA networks were constructed, from which we found that lncRNAs MSTRG.11484.2, MSTRG.32014.1 and MSTRG.29012.1 regulated at least three immune-related genes (ifih1, dhx58 and irf3) through PC-5p-43254_34, PC-3p-28352_70 and bta-miR-11987_L-1R-1_1ss8TA, and tap2 was regulated by two circRNAs (circRNA5279 and circRNA5277) by oni-mir-124a-2-p5_1ss13GA. The findings expand our understanding of the innate immune system of rainbow trout, and lay the foundation for further study of immune mechanisms and disease resistance breeding.

Analysis of yellow mutant rainbow trout transcriptomes at different developmental stages reveals dynamic regulation of skin pigmentation genes.

Yellow mutant rainbow trout (YR), an economically important aquaculture species, is popular among consumers due to its excellent meat quality and attractive appearance. Skin color is a key economic trait for YR, but little is known about the molecular mechanism of skin color development. In this study, YR skin transcriptomes were analyzed to explore temporal expression patterns of pigmentation-related genes in three different stages of skin color development. In total, 16,590, 16,682, and 5619 genes were differentially expressed between fish at 1 day post-hatching (YR1d) and YR45d, YR1d and YR90d, and YR45d and YR90d. Numerous differentially expressed genes (DEGs) associated with pigmentation were identified, and almost all of them involved in pteridine and carotenoid synthesis were significantly upregulated in YR45d and YR90d compared to YR1d, including GCH1, PTS, QDPR, CSFIR1, SLC2A11, SCARB1, DGAT2, PNPLA2, APOD, and BCO2. Interestingly, many DEGs enriched in melanin synthesis pathways were also significantly upregulated, including melanogenesis (MITF, MC1R, SLC45A2, OCA2, and GPR143), tyrosine metabolism (TYR, TYRP1, and DCT), and MAPK signaling (KITA) pathways. Using short time-series expression miner, we identified eight differential gene expression pattern profiles, and DEGs in profile 7 were associated with skin pigmentation. Protein-protein interaction network analysis showed that two modules were related to xanthophores and melanophores. In addition, 1,812,329 simple sequence repeats and 2,011,334 single-nucleotide polymorphisms were discovered. The results enhance our understanding of the molecular mechanism underlying skin pigmentation in YR, and could accelerate the molecular breeding of fish species with valuable skin color traits and will likely be highly informative for developing new therapeutic approaches to treat pigmentation disorders and melanoma.

Integrative mRNA-miRNA interaction analysis reveals the molecular mechanism of skin color variation between wild-type and yellow mutant rainbow trout (Oncorhynchus mykiss).

Rainbow trout (Oncorhynchus mykiss) is an important economic fish in China. Skin color affects the economic value of trout. However, the molecular mechanism of the skin color variation between wild-type (WR) and yellow mutant rainbow trout (YR) is unclear. We sequenced mRNAs and miRNAs of dorsal skin to identify key color variation-associated mRNAs and miRNAs between WR and YR. Overall, 2060 out of 3625 differentially expressed genes were upregulated in YR, and 196 out of 275 differentially expressed miRNAs were downregulated in WR. We identified three key YR-upregulated genes related to the formation of xanthophores (GCH1, SLC2A11, and SOX10). Interestingly, several genes related to melanogenesis (TYR, TYRP1, TYRP2, MC1R, MITF, PMEL, SLC45A2, and OCA2) were downregulated in WR. Integrated analysis identified five miRNAs that target at least two skin color-related genes (miR-495-y, miR-543-y, miR-665-z, miR-433-y, and miR-382-x). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses of target genes identified noncoding RNA metabolic process as the most significantly enriched GO term, and several metabolic pathways associated with skin color were enriched significantly, such as tyrosine metabolism, histidine metabolism, and vitamin B6 metabolism. Quantitative real-time PCR of selected mRNAs and miRNAs validated the reliability of the integrated analysis. This study provides in-depth insights into the molecular mechanism of skin color variation between WR and YR, which will accelerate the genetic selection and breeding of rainbow trout with consumer-favored traits.