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MiRNAs-Modulation of Nrf2 Signaling Networks in Regulation Oxidative Stress of Chinese Perch Skeletal Muscle After Fasting Treatment

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Abstract

Nrf2 is an important transcription factor involved in the antioxidant response and is widely expressed in animal tissues. The function of Nrf2 is regulated by its negative regulator Keap1 by inducing its cytoplasmic degradation. Recent studies have suggested that Nrf2 is also regulated post-transcriptionally via miRNAs. However, to date, how miRNAs regulate Nrf2 in fish skeletal muscles is unknown. In this study, the full-length cDNAs with 2398 bp of the Nrf2 was firstly cloned by SMART RACE amplification tools from Chinese perch. The Nrf2 gene structure and its 3’-UTR region for possible miRNA binding sites, as well as its spatial expression profile were assayed. Then, we employed TargetScan Fish tool MiRNAnome to predict putative sites for five miRNAs including miR-181a-5p, MiR-194a, MiR-216a, miR-459-5p, and miR-724. Using qRT-PCR assay, we found that Nrf2 mRNA levels have negative correlation with all five miRNAs expression in muscle of nutritionally deprived fish, and that ectopic expression of miR-181a-5p alone reduces Nrf2 mRNA levels. Luciferase reporter assay in a heterologous cell system revealed that each of the five miRNAs reduced Nrf2 expression, suggesting a direct regulatory mechanism. Moreover, the miR-181a-5p suppression using specific antagomir led to a significant increase in Nrf2 expression in vivo. At the same time, the expression levels of the antioxidant enzymes CAT, ZnSOD, GPx, GSTA, and GSTA genes increased significantly after injecting miR-181a-5p antagomir. Taken together, these findings provide evidence that miRNAs are involved in the Nrf2 signaling networks in regulation of oxidative stress in fish, at least in Chinese perch muscle.

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Acknowledgments

We thank Dr. NilliZmora in the University of Maryland reviewed and revised the manuscript.

Funding

This study was supported by the National Natural Science Foundation of China (Nos. 31820103016 and 31230076).

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Authors and Affiliations

  1. State Key laboratory of Chemo/Biosening and Chemomertrics, College of Biology, Hunan University, Changsha, 410082, People’s Republic of China

    Ping Wu, Xinhong Guo & Xuanming Liu

  2. Hunan Provincial Key Laboratory of Nutrition and Quality Control of Aquatic Animals, Department of Biological and Environmental Engineering, Changsha University, Changsha, 410022, China

    Ping Wu, Lin Chen, Jia Cheng, Yaxiong Pan, Wuying Chu & Jianshe Zhang

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  1. Ping Wu
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Correspondence to Jianshe Zhang or Xuanming Liu.

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This study was conducted following the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health of Hunan University, China. All procedures for fish handling were performed after fish were anesthetized using MS-222 (3-aminobenzoic acid ethyl ester methane sulfonate).

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Wu, P., Chen, L., Cheng, J. et al. MiRNAs-Modulation of Nrf2 Signaling Networks in Regulation Oxidative Stress of Chinese Perch Skeletal Muscle After Fasting Treatment. Mar Biotechnol 22, 620–630 (2020). https://doi.org/10.1007/s10126-020-09982-3

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