缩宫素受体主要存在形式的研究

A Study on Main Existence Form of Oxytocin Receptor

目的研究缩宫素受体(oxytocin receptor,OTR)在体外培养细胞中及动物组织中的主要存在形式,为进一步研究OTR多聚化的部位及多聚化在OTR功能调节中的作用提供依据。方法取体外培养的中国仓鼠(CHO)细胞中表达的OTR及大鼠不同的组织,分别用anti-OTR抗体进行免疫沉淀及免疫印迹,观察得到的OTR条带的情况;分别在样品中加入还原剂DTT,再进行免疫印迹,观察得到OTR条带的大小与数量。结果在体外培养的细胞中检测到OTR的多聚体,在大鼠的组织中同样也检测到OTR的多聚体。表明在动物体内,OTR也是以多聚体形式存在,跟体外细胞表达的OTR形成的多聚体不同的是,在动物体内的OTR多聚体主要以四聚体为主,而二聚体、三聚体量很少,特别是三聚体,几乎没有。OTR的多聚体可被还原剂DTT还原为单体形式。结论 OTR形成的多聚体是一种同源多聚体,此种多聚体中的单体是由共价键连接,而这种共价键最常见的是半胱氨酸残基间形成的二硫键,因此推测半胱氨酸残基是OTR多聚体形成的关键残基。

Objective To investigate the main existence form of the oxytocin receptor（OTR） in vitro culture cells and zoogenic tissues for further exploring the multimerization position of OTR and its adjustment in function of OTR.Methods The expression of OTR in Chinese hamsters cells and different tissues in rats were detected by immunoprecipitation and immunoblotting with Anti-OTR antibody.OTR strap was obtained and observed.The samples were detected for immunoblotting after DDT was added,and the size and number of OTR strap were observed.Results Multimer of OTR was found in both Chinese hamsters cells and rat tissues.The result showed that the main existence form in zoogenic tissues of OTR was multimer.Different from the OTR in CHO cells,the multimers in rat cells were mainly tetramers,but there were fewer dimerides and trimers,or even no trimers at all.The multimers could be reverted to monomers by DTT.Conclusion The monomers of OTR were homologous,and the monomers were connected by covalent bonds,which are mainly disulfide bonds formulated by Cys residue.So it is estimated that the disulfide bonds are essential in multimerization of OTR.