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JUN promotes chicken female differentiation by inhibiting Smad2

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Abstract

The sex determination and control of poultry is a key problem in production and scientific research despite few studies on regulatory factors, especially transcription factors in sex determination. In the early stage of this study, high-throughput sequencing was used to screen the differentially expressed gene JUN in male and female embryonic stem cells (ESCs) and primordial germ cells (PGCs). The qRT-PCR discovered that the JUN gene significantly increased from embryonic days (E) 2.5 later in chicken embryo development, and the female gonad expression was much higher than that of the male after E14.5. Lentivirus shRNA-JUN, shRNA-Smad2 interference, and OE-JUN overexpression vectors were successfully constructed. After interfering with JUN in vivo, male characteristics appeared in ZW embryonic gonads at E18.5. Meanwhile, the male-specific genes DMRT1 and Sox9 were upregulated, the female-specific genes FOXL2, ESR1, and CYP19A1 were downregulated, and the estradiol in the gonads was significantly decreased. The situation was reversed after the overexpression of JUN, ZZ chicken embryo developed into female sexual characteristics. The double luciferase report has found that the Smad2 promoter activity was significantly upregulated after interference with JUN, and significantly increased after the deletion of the JUN binding site. After the injection of the Smad2-shRNA vector into the blood vessel in vivo, it was discovered that DMRT1 and Sox9 of ZW embryos at E18.5 were downregulated, FOXL2 and CYP19A1 were significantly upregulated, and the gonads show femininity. In conclusion, this study proves that JUN is a key regulator in the process of chicken female sex differentiation, which can inhibit the transcription of Smad2 and promote the synthesis of estradiol, and participate in the process of chicken sex differentiation. This study lays a foundation for the analysis of the molecular mechanism of chicken sex determination and the development of poultry sex control technology.

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Acknowledgements

We would like to thank the research farm of the poultry research institute of the Chinese Academy of Agricultural Science for providing the Rugao yellow chicken eggs. This work was supported by the National Natural Science Foundation of China (31772582, 31972547), Key Research and Development Program(2017YFE0108000), High Level Talents Support Program of Yangzhou University, School Assistant of Jiangsu Graduate Practice Innovation Program(XSJCX19_100), albeit the funders had no role in study design, data collection, analysis, decision to publish, or preparation of the manuscript.

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Authors and Affiliations

  1. College of Animal Science and Technology, Jiangsu Province Key Laboratory of Animal Breeding and Molecular Design, Yangzhou University, 88 South University Ave, Yangzhou, 225009, Jiangsu, China

    Ming Zhang, Chen Zhang, Xiang Shi, Jancheng Li, Jingyi Jiang, Chen Chen, Yani Zhang, Guohong Chen, Bichun Li & Qisheng Zuo

  2. Department of Hematology, The People’s Hospital of Taizhou, Taizhou, Jiangsu, China

    Pei Xu

  3. Institute of Translational Medicine, Medical College, Yangzhou University, Yangzhou, 225009, Jiangsu, China

    Xiaolin Sun

Authors
  1. Ming Zhang
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  2. Pei Xu
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  3. Xiaolin Sun
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  4. Chen Zhang
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  5. Xiang Shi
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  6. Jancheng Li
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  8. Chen Chen
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  9. Yani Zhang
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  10. Guohong Chen
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  11. Bichun Li
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  12. Qisheng Zuo
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Contributions

LBC conceived and designed this study. ZM performed the experiments. ZM, ZC, XP, and SX collected and analyzed the data. LJC, JJY, and CC provided the reagents, materials, and analysis tools used in this study. ZM wrote the paper. SXL, ZQS, CGH, and ZYN revised the paper.

Corresponding author

Correspondence to Bichun Li.

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The authors declare that they have no competing interests.

Ethical approval

All procedures involving the care and use of animals conformed to the U.S. National Institute of Health guidelines (NIH Pub. No. 85-23, revised 1996) and were approved by the Laboratory Animal Management and Experimental Animal Ethics Committee of Yangzhou University.

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Zhang, M., Xu, P., Sun, X. et al. JUN promotes chicken female differentiation by inhibiting Smad2. Cytotechnology 73, 101–113 (2021). https://doi.org/10.1007/s10616-020-00447-y

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  • DOI: https://doi.org/10.1007/s10616-020-00447-y

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