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circRNA-miRNA-mRNA network analysis to explore the pathogenesis of abnormal spermatogenesis due to aberrant m6A methylation

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Abstract

Many studies have shown that circRNAs and miRNAs play important roles in many different life processes. However, the function of circRNAs in spermatogenesis remains unknown. Here, we aimed to explore the mechanisms whereby circRNA-miRNAs-mRNAs regulate abnormal m6A methylation in GC-1spg spermatogonia. We first reduced m6A methylation in GC-1spg whole cells after knocking down the m6A methyltransferase enzyme, METTL3. Then, we performed circRNA- and miRNA-seq on GC-1spg cells with low m6A methylation and identified 48 and 50 differentially expressed circRNAs and miRNAs, respectively. We also predicted the targets of the differentially expressed miRNAs by using Miranda software and further constructed the differentially expressed circRNA-differentially expressed miRNA-mRNA ceRNA network. GO analysis was performed on the differentially expressed circRNAs and miRNA-targeted mRNAs, and an interaction network between the proteins of interest was constructed using Cytoscape. The final GO analysis showed that the target mRNAs were involved in sperm formation. Therefore, a PPI network was subsequently constructed and 2 hub genes (H2afx and Dnmt3a) were identified. In this study, we constructed a ceRNA network and explored the regulatory roles of circRNAs and miRNAs in the pathogenesis of abnormal spermatogenesis caused by low levels of methylated m6A. Also, we identified two pivotal genes that may be key factors in infertility caused by abnormal m6A methylation. This may provide some ideas for the treatment of infertility resulting from abnormal spermatogenesis.

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Acknowledgements

The authors would like to thank Dr Dev Sooranna of Imperial College in London for editing the manuscript.

Funding

This work was supported by grants from the Guangxi Natural Science Foundation (No. 2020GXNSFBA297148 and AD20159062), the National Natural Science Foundation of China (No. 32102514, 31972996 and U20A2051), the Guangxi Science and Technology Major Project (No. GuiKe AA22068099), and the Bama County Program for Talents in Science and Technology, Guangxi, China (No. 202101265 and 202100174).

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Authors and Affiliations

  1. State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources, College of Animal Science and Technology, Guangxi University, Nanning, 530004, China

    Dandan Zhong, Liyin Zhang, Kongwei Huang, Mengjie Chen, Yaling Chen, Qingyou Liu, Deshun Shi & Hui Li

  2. Shaanxi Key Laboratory of Molecular Biology for Agriculture, College of Animal Science and Technology, Northwest A&F University, Yangling, 712100, China

    Hui Li

  3. Guangdong Provincial Key Laboratory of Animal Molecular Design and Precise Breeding, School of Life Science and Engineering, Foshan University, Foshan, 528225, China

    Qingyou Liu

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  1. Dandan Zhong
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Correspondence to Hui Li.

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Dandan Zhong, Liyin Zhang, and Kongwei Huang were responsible for the execution of the experiments and drawing diagrams. Dandan Zhong and Kongwei Huang participated in the writing of the manuscript. Kongwei Huang analyzed the sequencing data. Mengjie Chen and Yaling Chen were responsible for the analysis of experimental data. Qingyou Liu and Deshun Shi were involved in the revision of the manuscript. Hui Li was responsible for the experimental design and guided the project. Dandan Zhong and Liyin Zhang contributed equally to this work. All the authors read and approved the final manuscript.

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Zhong, D., Zhang, L., Huang, K. et al. circRNA-miRNA-mRNA network analysis to explore the pathogenesis of abnormal spermatogenesis due to aberrant m6A methylation. Cell Tissue Res 392, 605–620 (2023). https://doi.org/10.1007/s00441-022-03725-7

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