China Animal Husbandry & Veterinary Medicine ›› 2025, Vol. 52 ›› Issue (8): 3527-3539.doi: 10.16431/j.cnki.1671-7236.2025.08.003

• Biotechnology • Previous Articles    

Transcriptional Regulation of VTN Gene Promoter in Sheep Rumen Epithelial Cells

ZHONG Bingqian1, SUBINUR Eli1, ZHU Aiwen2, SUN Yan2, WANG Yutao1, YAN Wei2   

  1. 1. Key Laboratory of Biological Resources and Ecology of Pamirs Plateau in Xinjiang Uygur Autonomous Region, College of Life and Geographic Sciences, Kashi University, Kashi 844000, China;
    2. School of Animal Science and Technology, Jiangsu Agri-Animal Husbandry Vocational College, Taizhou 225300, China
  • Received:2024-11-04 Published:2025-08-02

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Abstract: 【Objective】 This study was aimed to investigate the transcriptional activity of the core promoter of vitronectin (VTN) gene in sheep rumem epithelial cells and the effects of CCCTC-binding factor (CTCF) gene overexpression on the expression of VTN and related genes. 【Method】 The core region of VTN gene promoter in sheep was predicted using online bioinformatics analysis software,and six pairs of promoter deletion fragment primers were designed.DNA was extracted from blood samples in Hu sheep to clone six promoter deletion fragments upstream of VTN gene CDS region.Luciferase recombinant plasmids L1-basic,L2-basic,L3-basic,L4-basic,L5-basic,and L6-basic were constructed and transfected into sheep rumen epithelial cells.Luciferase reporter gene assays were conducted to verify the location of promoter activity regions.The binding sites between transcription factor CTCF and VTN gene promoter were predicted using online bioinformatics analysis software.RNA was extracted from sheep rumen epithelial cells for reverse transcription,and the CDS region of CTCF gene in sheep was cloned to construct CTCF gene overexpression vector.The vector was co-transfected with VTN gene promoter recombinant plasmids L1-basic,L2-basic,L3-basic,and L4-basic into sheep rumen epithelial cells,with single plasmid transfection as control groups.Luciferase reporter gene assays were performed to determine the core region where transcription factor CTCF binds to VTN gene promoter.Real-time quantitative PCR was used to analyze the relative expression of CTCF,VTN,C-MYC,GLUT1,and SP1 genes in CTCF gene overexpression transfection groups. 【Result】 Six recombinant luciferase reporter vectors in the promoter region of VTN gene in sheep were successfully constructed.The results of bioinformatics prediction and luciferase reporter gene assays indicated that the core promoter of VTN gene might be located at ―933/―440 bp,and transcription factor CTCF potentially interacted with the core region of VTN gene promoter.After CTCF gene overexpression,compared with control group,the relative expression of VTN,C-MYC,and GLUT1 genes were significantly decreased (P＜0.05),while the expression of SP1 gene showed no significant difference (P＞0.05). 【Conclusion】 In sheep rumen epithelial cells,transcription factor CTCF might predominantly bind to the promoter region (―439/+26 bp) and core region (―933/―440 bp) to influence VTN gene expression.CTCF gene overexpression affected the transcription of VTN,C-MYC,and GLUT1 genes,suggesting that CTCF gene potentially influenced the growth,proliferation,and metabolism of sheep rumen epithelial cells.

Key words: sheep; VTN gene; core promoter; rumem epithelial cells; CTCF gene

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