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表皮生长因子受体(epidermal growth factor receptor, EGFR)是酪氨酸激酶受体家族成员之一,不仅参与细胞增殖、生长和凋亡等多种生命活动,也可调节哺乳动物的乳腺发育及泌乳维持,但对绵羊EGFR基因的序列特征及组织表达情况鲜有报道。本试验以高泌乳量的小尾寒羊(泌乳高峰期和空怀期)及低泌乳量的甘肃高山细毛羊(泌乳高峰期)母羊为研究对象,利用RT-PCR、克隆及测序技术获得绵羊EGFR基因完整的CDS区,分析了EGFR蛋白的结构特征及理化性质,利用RT-qPCR技术研究了基因的组织表达情况。结果表明,绵羊EGFR基因CDS区全长为3 627 bp,编码1 208个氨基酸。绵羊EGFR的氨基酸序列在各物种间较保守,与黄牛EGFR的氨基酸序列同源性最高。EGFR为跨膜蛋白,包含111个磷酸化位点,二级结构以α螺旋和无规则卷曲为主。网络互作分析表明EGFR蛋白与肝素结合表皮生长因子(HB-EGF)、表皮调节素(EREG)、双调蛋白(AREG)及生长因子受体结合蛋白2 (GRB2)结合发挥作用。EGFR主要参与MAPK,PI3K/AKT,JAK/STAT及Wnt信号通路,从而参与了动物的乳腺发育及泌乳功能的调节。RT-qPCR结果表明,绵羊EGFR基因的表达具有组织特异性、时空特异性和品种特异性。该基因在所研究的8个组织中均表达,但在肾脏、卵巢、肝脏、乳腺和肺脏组织中的表达量较高;在小尾寒羊的乳腺组织中,该基因在空怀期的表达量显著高于泌乳高峰期的(P<0.05);在泌乳高峰期的乳腺组织中,该基因在小尾寒羊中的表达量高于甘肃高山细毛羊的。本试验为深入研究绵羊EGFR基因的泌乳生物学功能提供了基础数据。
Abstract：Epidermal growth factor receptor(EGFR) is a member of the tyrosine kinase receptor family. EGFR plays an important role in cell proliferation, growth and apoptosis, and regulates mammalian mammary gland development and lactation. However, there are few reports on the sequence characteristics and tissue expression of sheep EGFR gene. Small-tailed Han sheep with high milk yield(peak lactation period and empty period) and Gansu alpine fine wool sheep with low milk yield(peak lactation period) were used in this experiment. The intact CDS region of sheep EGFR gene was obtained by RT-PCR, cloning and sequencing. The structural characteristics and physicochemical properties of EGFR protein were analyzed. RT-qPCR was used to detect the expression of EGFR gene in tissues.The results showed that the CDS region of sheep EGFR gene was 3 627 bp, encoding 1 208 amino acids. The amino acid sequence of sheep EGFR was conserved among different species, it had the highest amino acid sequence homology with cattle EGFR. EGFR was a transmembrane protein with 111 phosphorylation sites. The secondary structure of EGFR was mainly composed of α-helix and random coil. String analysis showed that EGFR protein interacted with heparin-binding epidermal growth factor(HB-EGF), epiregulin(EREG), amphiregulin(AREG) and growth factor receptor binding protein 2(GRB2). The EGFR was mainly showed in MAPK, PI3 K/AKT, JAK/STAT and Wnt signaling pathways, which participated in the regulation of animal mammary gland development and lactation. RT-qPCR results showed that the expression of EGFR gene was tissue-specific, space-time specific and breed-specific. The gene was expressed in the 8 tissues studied, but showed higher eexpression in kidney, ovary, liver, mammary gland and lungs. In the mammary gland tissue of Small Tail Han sheep, the expression of EGFR gene in the empty period was significantly higher than that in the peak of lactation(P<0.05). In the mammary gland tissue at the peak of lactation, the expression of EGFR gene in small-tailed Han sheep was higher than that in Gansu alpine fine-wool sheep. This study provided basic data for in-depth study of the lactational biological function of the sheep EGFR gene.
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基本信息:
DOI：10.13417/j.gab.040.001463
中图分类号:S826
引用信息:
[1]沈继源,王继卿,郝志云,等.绵羊EGFR基因的克隆、表达及序列分析[J].基因组学与应用生物学,2021,40(04):1463-1470.DOI:10.13417/j.gab.040.001463.
基金信息:
国家自然科学基金项目(31860635); 甘肃农业大学“伏羲青年英才培育计划”项目(Gaufx-02Y02)和甘肃农业大学自列课题(GSAU-ZL-2015-033)共同资助
2021-04-25
2021-04-25