文昌鱼tropomyosin基因的克隆、进化分析及其胚胎发育与成体中的表达模式
Phylogenetic analysis and expression patterns of tropomyosin in amphioxus
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摘要: Tropomyosin是一种分布广泛而且在进化上十分保守的蛋白, 是肌肉形成和收缩过程中重要的调节蛋白质。通过RT-PCR和RACE技术得到文昌鱼tropomyosin基因全长, 编码一个含284个氨基酸残基的蛋白质, 将文昌鱼Tropomyosin和在其他物种中的同源物进行比对建树, 发现其在功能域上高度保守并且只有一个拷贝, 符合动物分类学中各物种的进化地位。胚胎整体原位杂交实验得知, tropomyosin在文昌鱼早期发育的表达,最早从原肠胚末期神经胚早期开始, 定位于分化中的中内胚层。到神经胚期, tropomyosin的表达出现在发育中的体节和脊索中。随着发育的进行, tropomyosin的表达稳定地集中在体节、脊索处。到72 h幼虫阶段, tropomyosin的表达仍然在肌节内。成体的切片原位杂交结果显示, tropomyosin在肌节中的表达大幅度下调, 而在神经管细胞、脊索和腮区腮瓣处仍然可以检测到明显的表达, 在外胚层和表皮内没有发现杂交信号。研究结果表明, tropomyosin的表达与文昌鱼肌节、肌肉以及神经索的发生相关, 参与文昌鱼胚胎躯体模式的构建, 而且在成体的生命活动中发挥重要作用。Abstract: In amphioxus, we found a mesoderm related gene, tropomyosin,which encodes a protein comprising 284 amino acid residues, sharing high identities with other known Tropomyosin proteins both in vertebrates and invertebrates. Phylogenetically, amphioxus Tropomyosin fell outside the invertebrate clade and was at the base of the vertebrate protein family clade, indicating that it may represent an independent branch. From the early neurula to the larva stage, whole-mount in situ hybridization and histological sections found transcripts of amphioxus tropomyosin gene. Weak tropomyosin expression was first detected in the wall of the archenteron at about 10 hours-post-fertilization neurula stage, while intense expression was revealed in the differentiating presumptive notochord and the muscle. Transcripts of tropomyosin were then expressed in the formed notochord and somites. Gene expression seemed to continue in these developing organs throughout the neurular stages and remained till 72-hours, during the early larval stages. In situ study still showed tropomyosin was also expressed in the neural tube, hepatic diverticulum, notochord and the spaces between myotomes in adult amphioxus. Our results indicated that tropomyosin may play an important role in both embryonic development and adult life.