唇䱻bmp2a基因克隆及其与肌间刺骨化相关性分析

MOLECULAR CHARACTERIZATION OF A BMP2A HOMOLOGUE IN BARBEL STEED (HEMIBARBUS LABEO) AND ITS INVOLVEMENT IN INTERMUSCULAR BONE DEVELOPMENT

  • 摘要: 骨形成蛋白(Bone morphogenetic protein, BMP)属于转化生长因子β(Transforming growth factor-beta, TGF-β)超家族。在骨形成蛋白家族中, BMP2属于分泌性多功能蛋白, 具有较强的诱导骨细胞形成的能力。为了探究BMP2与肌间刺骨化的相关性,研究通过转录组测序和RT-PCR获得了唇䱻(Hemibarbus labeo)bmp2a基因cDNA序列。氨基酸序列比对结果显示, 唇䱻BMP2A和其他鱼类BMP2保守性较高, 都具有一个高度保守的TGFβ结构域, 在该结构域中存在一个N-糖基化位点。此外, 包括唇䱻BMP2A在内, 鱼类BMP2都具有7个保守的半胱氨酸残基。系统进化树分析表明, 唇䱻BMP2A与团头鲂(Megalobrama amblycephala)BMP2A最为接近。通过荧光定量PCR检测发现唇䱻bmp2a基因在所有被检测的组织中均有表达, bmp2a基因在鳃中的表达量最高, 肝脏和肌肉中相对较高, 在心脏中表达量最低。免疫组化结果显示BMP2A分布于肌肉肌隔中。此外, bmp2a基因的表达与肌间刺骨化时机相吻合。综上, 唇䱻BMP2A与肌间刺骨化存在较高的相关性。研究结果将为进一步调查鱼类BMP2功能以及鱼类肌间刺形成分子机制提供理论依据。

     

    Abstract: Bone morphogenetic protein (BMP) belongs to the transforming growth factor-beta (TGF-β) superfamily, and BMP2 is a secreted multifunctional protein with strong ability to induce bone formation. To explore the correlation between BMP2 and intermuscular bone development, the bmp2a cDNA of barbel steed (Hemibarbus labeo) was cloned and sequenced. The amino acid sequence alignment showed that the barbel steed BMP2A is highly conserved with other fishes, and they all have the highly conserved TGFβ domain in which an N-glycosylation site exists. In addition, fish BMP2 protein, including barbel steed BMP2A, have seven conserved cysteine residues. Phylogenetic tree analysis indicated that barbel steed BMP2A were closely related to that of Wuchang bream (Megalobrama amblycephala) BMP2A. The expression of barbel steed bmp2a transcripts was detected in all tested tissues, with the highest mRNA level in the gill, followed by the liver and muscle, and the lowest in the heart. Immunohistochemistry results showed that BMP2A protein was distributed in the myosepta. In addition, the expression of the bmp2a gene coincides with the development of intermuscular ossification. In summary, the high correlation between bmp2a gene and intermuscular ossification provides basic data for further investigation of the function of bmp2 gene and its regulation role in intermuscular bone development.

     

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