Li Xuening, Hu Tingting, Li Ruiwen, Li Yanyan, Lin Yaqiu, Wang Yong, Liu Wei, Wang Youli
Key Laboratory of Qinghai-Tibetan Plateau Animal Genetic Resource Reservation and Utilization of Education Ministry, Southwest Minzu University, Chengdu, China.
Key Laboratory of Qinghai-Tibetan Plateau Animal Genetic Resource Reservation and Exploitation of Sichuan Province, Southwest Minzu University, Chengdu, China.
Anim Biosci. 2025 May;38(5):873-883. doi: 10.5713/ab.24.0248. Epub 2024 Oct 28.
The aim of this study was to obtain goat CRTC2 gene sequence and elucidate its biological properties, and further study the impact of overexpression and interference of CRTC2 on the cell differentiation of goat subcutaneous precursor adipocytes.
The sequence of goat CRTC2 was cloned by reverse transcription (RT)-polymerase chain reaction (PCR) and its molecular characterization was analyzed. The expression of CRTC2 gene in goat tissues and subcutaneous precursor adipocytes differentiated from 0 to 120 h was examined by quantitative real-time PCR (qRT-PCR). The effects of CRTC2 on the subcutaneous precursor adipocyte differentiation were investigated by using liposome transfection, Bodipy, Oil Red O staining and qPCR.
The results showed that the cloned goat CRTC2 gene was 2363 bp long (coding sequence [CDS] 2082 bp), encoding 693 amino acids. The relative expression levels of CRTC2 gene were highest in liver and then in kidney (p<0.05). During differentiation, the highest expression of CRTC2 in subcutaneous precursor adipocytes was observed at 120 of differentiating (p<0.01). In addition, we found that overexpression of CRTC2 significantly increased the expression of lipid metabolism-related genes (C/EBPα, C/EBPβ, PPARγ, DGAT1, DGAT2, ACC, FASN, SREBP1, AP2, LPL, ATGL) and promoted lipid accumulation. We then chemically synthesized goat CRTC2 small interfering RNA and transfected it into goat subcutaneous precursor adipocytes. The results revealed that SiRNA-mediated interference with CRTC2 significantly inhibited its differentiation and suppressed lipid droplet aggregation.
So, this study indicates that CRTC2 is a positive regulator that promoting cell differentiation of subcutaneous adipocyte in goats, which lays the foundation for an in-depth study of the role of CRTC2 in lipid deposition in goats.
本研究旨在获取山羊CRTC2基因序列，阐明其生物学特性，并进一步研究CRTC2过表达和干扰对山羊皮下前体脂肪细胞分化的影响。
通过逆转录（RT）-聚合酶链反应（PCR）克隆山羊CRTC2序列，并分析其分子特征。采用定量实时PCR（qRT-PCR）检测CRTC2基因在山羊组织及0至120小时分化的皮下前体脂肪细胞中的表达。利用脂质体转染、Bodipy、油红O染色和qPCR研究CRTC2对皮下前体脂肪细胞分化的影响。
结果显示，克隆的山羊CRTC2基因长2363 bp（编码序列[CDS] 2082 bp），编码693个氨基酸。CRTC2基因相对表达水平在肝脏中最高，其次是肾脏（p<0.05）。在分化过程中，皮下前体脂肪细胞中CRTC2在分化120小时时表达最高（p<0.01）。此外，我们发现CRTC2过表达显著增加脂质代谢相关基因（C/EBPα、C/EBPβ、PPARγ、DGAT1、DGAT2、ACC、FASN、SREBP1、AP2、LPL、ATGL）的表达并促进脂质积累。然后我们化学合成山羊CRTC2小干扰RNA并将其转染到山羊皮下前体脂肪细胞中。结果表明，SiRNA介导的对CRTC2的干扰显著抑制其分化并抑制脂滴聚集。
因此，本研究表明CRTC2是促进山羊皮下脂肪细胞分化的正调控因子，为深入研究CRTC2在山羊脂质沉积中的作用奠定了基础。
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