华北农学报 ›› 2025, Vol. 40 ›› Issue (3): 225-233. doi: 10.7668/hbnxb.20195346

所属专题: 畜牧

• 畜牧·水产·兽医 • 上一篇    下一篇

牦牛MAP3K7基因克隆及其与皮下前体脂肪细胞分化关系分析

蓝湘一, 何洋洋, 张明, 黄慈, 王海波, 王会, 柴志欣, 钟金城   

  1. 西南民族大学 青藏高原动物遗传资源保护与利用四川省重点实验室,青藏高原动物遗传资源保护与利用教育部重点实验室,四川 成都 610041
  • 收稿日期:2024-08-07 出版日期:2025-07-04
  • 通讯作者:
    钟金城(1963—),男,云南绿春人,教授,博士,博士生导师,主要从事动物遗传育种与繁殖研究。
  • 作者简介:

    蓝湘一(2000—),女,广西宜州人,硕士,主要从事分子遗传学研究。

  • 基金资助:
    国家自然科学基金项目(32372849); 四川省科技计划项目(2024NSFSC0301); 青海省科技计划项目(2022-NK-110); 西南民族大学中央高校基本科研业务费专项资金(2023NYXXS097)

Cloning of Yak MAP3K7 Gene and Its Relationship with the Differentiation of Subcutaneous Preadipocytes

LAN Xiangyi, HE Yangyang, ZHANG Ming, HUANG Ci, WANG Haibo, WANG Hui, CHAI Zhixin, ZHONG Jincheng   

  1. Qinghai-Tibetan Plateau Animal Genetic Resource Reservation and Utilization Key Laboratory of Sichuan Province,Key Laboratory of Qinghai-Tibetan Plateau Animal Genetic Resource Reservation and Utilization of Ministry of Educacation, Southwest Minzu University,Chengdu 610041,China
  • Received:2024-08-07 Published:2025-07-04

摘要:

MAP3K7是MAP3K家族中的一员,在脂肪分化、炎症和肿瘤发生过程中发挥着重要作用,但其与牦牛脂肪分化的关系未见报道,并且其基因序列未知,制约了相关功能研究。为了获得牦牛MAP3K7 基因编码区 (CDS) 保守区域序列,明确其表达特征,了解其在牦牛不同组织中的表达水平及其与牦牛皮下前体脂肪细胞分化的相关性,利用RT-PCR技术,克隆牦牛 MAP3K7 基因并利用生物信息学方法分析生物学特性;利用实时荧光定量 PCR (qPCR) 技术检测MAP3K7 在牦牛心、肝、脾等10个组织中的表达情况,与不同分化阶段牦牛皮下脂肪细胞中MAP3K7基因和脂分化相关基因的表达情况,同时结合油红O染色,明确MAP3K7 基因对牦牛皮下前体脂肪细胞分化的影响。结果表明:克隆所得牦牛MAP3K7基因CDS区长度为1 740 bp,编码579个氨基酸,与野牦牛MAP3K7 预测序列CDS区比对发现第967位 A>G,导致第323位氨基酸Thr>Ala的突变。理化性质分析发现,该蛋白为整体带负电的亲水酸性蛋白,具有112个氨基酸磷酸化位点,其氨基酸组成中,占比最高的是丝氨酸;二级结构预测显示,其无规卷曲含量最高,可能与TAB3等蛋白质间存在相互作用关系。核苷酸同源性比对与进化树结果表明,牦牛MAP3K7在反刍动物间具有较高的保守性,与野牦牛同源性最高且进化关系最近。组织定量检测发现,MAP3K7在牦牛皮下脂肪和内脏脂肪中的表达量相对较低,在牦牛皮下前体脂肪细胞诱导分化过程中,MAP3K7表达量随着成熟脂肪细胞的增多先上调,至分化到一定程度后下调。综上,MAP3K7基因在牦牛皮下前体脂肪细胞分化前期高表达以促进分化,但当分化到一定程度时,表达下调抑制进一步分化。

关键词: 牦牛, MAP3K7, 基因表达, 皮下脂肪, 脂肪分化

Abstract:

MAP3K7 is a member of the MAP3K family and plays an important role in fat differentiation,inflammation and tumorigenesis.However,its relationship with yak fat differentiation has not been reported,and its gene sequence is unknown,which restricts the related functional research.In order to obtain the conserved region sequence of yak MAP3K7 gene coding region (CDS),clarify its expression characteristics,and understand its expression level in different tissues of yak and its correlation with subcutaneous preadipocyte differentiation of yak,the MAP3K7 gene of yak was cloned by RT-PCR and the biological characteristics were analyzed by bioinformatics methods.Real-time Fluorescence Quantitative PCR (qPCR) was used to detect the expression of MAP3K7 in 10 tissues of yak heart,liver,spleen,etc,and the expression of MAP3K7 gene and lipid differentiation-related genes in yak subcutaneous adipocytes at different differentiation stages.At the same time,combined with oil red O staining,the effect of MAP3K7 gene on the differentiation of yak subcutaneous preadipocytes was clarified.The results showed that the length of the CDS region of the cloned yak MAP3K7 gene was 1 740 bp,encoding 579 amino acids.Compared with the CDS region of the wild yak MAP3K7 predicted sequence,it was found that A>G at position 967,resulting in a mutation of Thr>Ala at position 323.The analysis of physical and chemical properties showed that the protein was a negatively charged hydrophilic acidic protein with 112 amino acid phosphorylation sites.Among its amino acid composition,serine accounted for the highest proportion.The secondary structure prediction showed that the content of random coil was the highest,which might interact with proteins such as TAB3.The results of nucleotide homology comparison and phylogenetic tree showed that yak MAP3K7 was highly conserved among ruminants,and had the highest homology and the closest evolutionary relationship with wild yak.Tissue quantitative detection showed that the expression of MAP3K7 in subcutaneous fat and visceral fat of yak was relatively low.During the differentiation of subcutaneous preadipocytes of yak,the expression of MAP3K7 was up-regulated with the increase of mature adipocytes,and then down-regulated after differentiation to a certain extent.In summary,MAP3K7 gene was highly expressed in the early stage of differentiation of yak subcutaneous preadipocytes to promote differentiation,but when the differentiation reached a certain level,the down-regulation of expression inhibited further differentiation.

Key words: Bos grunniens, MAP3K7, Gene expression, Subcutaneous fat, Adipose differentiation

中图分类号: 

引用本文

蓝湘一, 何洋洋, 张明, 黄慈, 王海波, 王会, 柴志欣, 钟金城. 牦牛MAP3K7基因克隆及其与皮下前体脂肪细胞分化关系分析[J]. 华北农学报, 2025, 40(3): 225-233. doi: 10.7668/hbnxb.20195346.

LAN Xiangyi, HE Yangyang, ZHANG Ming, HUANG Ci, WANG Haibo, WANG Hui, CHAI Zhixin, ZHONG Jincheng. Cloning of Yak MAP3K7 Gene and Its Relationship with the Differentiation of Subcutaneous Preadipocytes[J]. Acta Agriculturae Boreali-Sinica, 2025, 40(3): 225-233. doi: 10.7668/hbnxb.20195346.