秦川牛MGP基因生物信息学分析及表达特性研究

doi:10.16431/j.cnki.1671-7236.2024.06.002

摘要/Abstract

摘要： 【目的】预测基质Gla蛋白(matrix Gla protein,MGP)的基本功能,揭示MGP基因在秦川牛中的组织表达规律以及在秦川牛前体脂肪细胞不同分化时期的表达特性,为进一步研究MGP基因对秦川牛脂肪发育调控的作用机制提供理论依据。【方法】运用生物信息学在线软件对多物种MGP蛋白进行相似性比对及系统进化树构建,并对牛MGP蛋白的理化性质、亚细胞定位等进行预测;使用油红O染色检验秦川牛前体脂肪细胞的分化效果;利用实时荧光定量PCR技术检测MGP基因在秦川牛不同组织中的表达情况,以及在前体脂肪细胞不同分化时期的表达特性。【结果】相似性比对结果显示,牛MGP蛋白与山羊的相似性最高(99%),与斑马鱼的相似性最低(31%)。系统进化树显示,牛和山羊的亲缘关系最近,与斑马鱼的亲缘关系最远。MGP基因编码103个氨基酸;理论等电点为9.27,偏碱性;总平均疏水指数为—0.630,为亲水性蛋白;含有Sec/SPⅠ类型的信号肽,无跨膜结构,属于分泌蛋白;存在5个丝氨酸磷酸化位点和4个酪氨酸磷酸化位点,具有包含γ-羧基谷氨酸(Gla)的GLA_domain;二级结构中α-螺旋占比最高(55.34%)。油红O染色结果确定了分离培养的前体脂肪细胞分化效果良好。实时荧光定量PCR结果显示,MGP基因在秦川牛心脏中的表达量显著高于其他组织(P＜0.05),在大肠和肾周脂肪中也有较高的表达水平,在小肠中的表达量显著低于其他组织(P＜0.05);MGP基因在前体脂肪细胞不同分化时期呈现先升高后降低的表达趋势,在前体脂肪细胞诱导分化第2天的表达量显著高于其他时间点(P＜0.05),第10天的表达量显著低于其他时间点(P＜0.05)。【结论】试验预测了MGP蛋白基本的结构和功能;MGP基因在秦川牛心脏组织中表达量最高,在小肠组织中表达量最低;MGP基因在前体脂肪细胞不同分化时期表现出先升高后降低的表达趋势。

关键词: 秦川牛; MGP基因; 组织表达; 前体脂肪细胞

Abstract: 【Objective】 This study was aimed to predict the fundamental functions of matrix Gla protein (MGP),reveal the tissue expression patterns of MGP gene in Qinchuan cattle and its expression characteristics during different stages of preadipocyte differentiation,so as to provide a theoretical foundation for studying the regulatory mechanisms of MGP gene in the adipose development of Qinchuan cattle.【Method】 The similarity alignment and phylogenetic tree construction of MGP protein in various species were conducted,and the physicochemical properties and subcellular localization of bovine MGP protein were predicted using bioinformatics online software.The differentiation effect of preadipocytes in Qinchuan cattle was examined using oil Red O staining.The expression of MGP gene in different tissues of Qinchuan cattle and its characteristics during different stages of preadipocyte differentiation were detected by Real-time quantitative PCR.【Result】 The similarity alignment results showed that the similarity of MGP protein was the highest (99%) between Bos taurus and Capra hircus,while the similarity was the lowest (31%) with Danio rerio.The phylogenetic tree revealed that Bos taurus and Capra hircus shared the closest relationship,whereas the relationship was the most distant with Danio rerio.MGP gene encoded 103 amino acids with a theoretical isoelectric point of 9.27,indicating alkalinity.MGP was considered a hydrophilic protein with a total average hydrophobicity index of ―0.630.MGP was classified as a secretory protein due to the presence of a Sec/SPⅠ-type signal peptide and the absence of transmembrane structures.Additionally,MGP protein included 5 serine and 4 threonine phosphorylation sites,and it possessed a GLA_domain containing γ-carboxyglutamic acid (Gla).The alpha helix constituted the highest percentage of the secondary structure at 55.34%.The oil Red O staining confirmed the well-differentiated state of isolated and cultured preadipocytes.Real-time quantitative PCR results showed that the expression of MGP gene in heart of Qinchuan cattle was significantly higher than that in other tissues (P＜0.05).Elevated expression were also observed in large intestine and perirenal fat,while the expression in small intestine was significantly lower than that in other tissues (P＜0.05).MGP gene exhibited a trend of initial upregulation followed by downregulation during different stages of preadipocyte differentiation,with significantly higher expression on the 2nd day of preadipocyte induction compared with other days (P＜0.05),and significantly lower expression on the 10th day compared with other days (P＜0.05).【Conclusion】 This study predicted the basic structure and function of MGP protein.The expression of MGP gene was the highest in heart of Qinchuan cattle and was the lowest in small intestine.MGP gene exhibited a trend of initial upregulation followed by downregulation during different stages of preadipocyte differentiation.

Key words: Qinchuan cattle; MGP gene; tissue expression; preadipocyte

中图分类号:

蒋蕾, 王玉鑫, 万媛, 昝林森, 王洪宝. 秦川牛MGP基因生物信息学分析及表达特性研究[J]. 中国畜牧兽医, 2024, 51(6): 2261-2272.

JIANG Lei, WANG Yuxin, WAN Yuan, ZAN Linsen, WANG Hongbao. Bioinformatics and Expression Characteristics Analysis of MGP Gene in Qinchuan Cattle[J]. China Animal Husbandry and Veterinary Medicine, 2024, 51(6): 2261-2272.

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