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FAS配体 FASL 在COS |
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罗振革 彭 虹 孔祥英 高杰英
摘 要 目的：获得FASL的真核细胞表达，并分析其功能。方法：将编码人FASL(FAS ligand)的全长cDNA插入真核细胞瞬时表达载体PSVL的SV40晚期启动子下游的XbaI位点，构建FASL的真核细胞表达载体PSVL-hFASL，用电击法将PSVL-hFASL转染COS-7细胞，转染后48 h，用Northern-blot法检测FASL的表达。结果：转染后COS-7细胞有FASL的mRNA的表达，在COS-7细胞上表达的重组FASL能诱导FAS阳性的U937细胞发生程序性死亡。结论：在COS-7细胞表面表达的重组FASL分子能用于进一步功能研究和FAS系统拮抗剂的筛选。
关键词 FASL 真核表达 程序性细胞死亡
中国图书分类号 R392.12
Expression of recombinant FAS ligand in COS-7 cells
LUO Zhen-Ge,PENG Hong,KONG Xiang-Ying et al.
Institute of Microbiology and Epidemiology,Academy of Military Medical Sciences,Beijing 100850
Abstract Objective：Obtain recombinant FASL molecules in COS-7 cell line and study the functions of FASL. Methods:The cDNA for human FAS ligand(FASL) was cloned into the downstream of SV40 late promoter of PSVL plasmid, which is a eukaryotic transient expressive vector. By electroporation, PSVL-hFASL was transfected into cos-7 cells.48 h after transfection,whole RNA of transfected cos-7 cells was prepared and analyzed by Northern-blot.Results:PSVL-hFASL transfected cells had the expression of human FAS ligand mRNA, PSVL mock transfected cells was negative. There is expression of FAS antigen on the surface of U937 cells. With U937 cell line as target, the activity of recombinant human FAS ligand (rhFASL) was assayed. After coculture of U937 cells with FASL transfected COS-7, the target cells appeared apoptosis phenomenom. Conclusion:Recombinant FasL on the surface of COS-7 cells can be used as functional study.
Key words FASL Eukaryotic expression Apoptosis
FAS是一个Ⅰ型跨膜受体分子,属于TNF/NGF受体超家族的成员,存在于多种细胞表面,FAS的配体（抗FAS抗体或其天然配体FASL）与FAS胞外区结合后，使其形成三聚体，通过FAS胞浆区中“death domain”间作用，传递死亡信号,引起靶细胞的程序性死亡［1］。FASL(FAS ligand)是FAS的天然配体,为与TNF同源的Ⅱ型穿膜分子，主要分布于活化的T细胞及一些免疫屏蔽器官［2，3］。FAS系统不仅参与免疫细胞发育,还能下调免疫反应,从而维持免疫平衡,介导CTL及NK细胞对靶细胞的杀伤作用［4，5］。最近还有人报道了FAS系统介导的CD4+的Th1细胞诱导B细胞凋亡的作用［6］。FAS系统还与自身免疫病、移植排斥、病毒感染和肿瘤发生有密切关系［7，8］。为了深入分析FAS系统的功能和作用机制及在某些疾病发生、发[1] [2] 下一页
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