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Anim Biosci > Volume 37(12); 2024 > Article
Animal Breeding and Genetics
Animal Bioscience 2024;37(12): 2066-2080.
https://doi.org/10.5713/ab.24.0081    Published online August 22, 2024.
N6-methyladenosine (m6A)-circHECA from secondary hair follicle of cashmere goats: identification, regulatory network and expression regulated potentially by methylation of its host gene promoter
Jincheng Shen1  , Taiyu Hui1  , Man Bai1  , Yixing Fan1  , Yubo Zhu1  , Qi Zhang1  , Ruqing Xu1  , Jialiang Zhang1  , Zeying Wang1  , Wenxin Zheng2,3  , Wenlin Bai1,* 
1College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Shenyang 110866, China
2State Key Laboratory for Herbivorous Livestock Genetic Improvement and Germplasm Innovation of Ministry of Science and Technology and Xinjiang Uygur Autonomous Region, Urumqi 830011, China
3Xinjiang Academy of Animal Sciences, Urumqi 830011, China
Correspondence:  Wenlin Bai, Tel: +86-24-8848-7156, Email: baiwenlin@syau.edu.cn
Received: 7 February 2024   • Revised: 2 April 2024   • Accepted: 11 May 2024
Abstract
Objective
The objective of this study was to identify the N6-methyladenosine (m6A)- circHECA molecule in secondary hair follicles (SHFs) of cashmere goats, and generate its potential regulatory network, as well as explore the potential relationship between transcriptional pattern of m6A-circHECA and promoter methylation of its host gene (HECA).
Methods
The validation of circHECA m6A sites was performed using methylation immunoprecipitation (Me-RIP) along with reverse transcription-quantitative polymerase chain reaction (RT-qPCR) technique. The nucleus and cytoplasm localizations of m6AcircHECA were performed using SHF stem cells of cashmere goats with RT-qPCR analysis. Based on in-silico analysis, the regulatory networks of m6A-circHECA were generated with related signal pathway enrichment. The methylation level of promoter region of m6A-circHECA host gene (HECA) was assessed by the bisulfite sequencing PCR (BSPPCR) technique.
Results
The m6A-circHECA was confirmed to contain four m6A modification sites including m6A-213, m6A-297, m6A-780, and m6A-927, and it was detected mainly in cytoplasm of the SHF stem cells of cashmere goats. The integrated regulatory network analysis showed directly or indirectly complex regulatory relationships between m6A-circHECA of cashmere goats and its potential target molecules: miRNAs, mRNAs, and proteins. The regulatory network and pathway enrichment indicated that m6A-circHECA might play multiple roles in the SHF physiology process of cashmere goats through directly or indirectly interacting or regulating its potential target molecules. A higher methylation level of promoter region of HECA gene in SHFs of cashmere goats might cause the lower expression of m6A-circHECA.
Conclusion
The m6A-circHECA might play multiple roles in SHF physiology process of cashmere goats through miRNA mediated pathways along with directly or indirectly interaction with its target proteins. The promoter methylation of m6A-circHECA host gene (HECA) most likely was implicated in its expression inhibition in SHFs of cashmere goats.
Keywords: Cashmere Goats; CircHECA; N6-Methyladenosine; Promoter Methylation; Regulatory Network; Transcriptional Pattern
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