CⅡTA M1-RNA对MHCⅡ类分子表达的抑制作用  

The inhibitory effect of anti-CⅡ TA M1-RNA on MHCⅡ expression in ECV304 cells

作　　者：郭荣[1] 何飞[2] 杜欣[1] 翁建宇[1] 陆泽生[1] 林伟[1] 

机构地区：[1]广东省人民医院血液科,广州510080 [2]广东省心血管病研究所心内科

出　　处：《中华微生物学和免疫学杂志》2007年第12期1071-1075,共5页Chinese Journal of Microbiology and Immunology

基　　金：广东省医学科学技术研究基金(A2005017);广东省自然科学基金(04003959;06020896);国家自然科学基金(30571771)

摘　　要：目的探讨MHCⅡ类分子转录激活因子(CⅡTA)的M1-RNA对细胞表面MHCⅡ类分子表达的抑制。方法M1-RNA是核糖核酸酶P的催化活性单位,设计并克隆针对CⅡTA第452、629位点的M1-RNA(分别为M1-452-GS、M1-629-GS)及其相应的CⅡTA靶基因,分别插入pUC19、pGEM-7zf (+)载体,进行细胞外切割活性筛选。将细胞外切割作用明显的M1-629-GS亚克隆入psNAV载体(psNAV-M1-629-GS,pA629)并稳定转染ECV304细胞株,流式细胞术检测经典的MHCⅡ(HLA-DR、-DP、-DQ)类抗原表达,RT-PCR检测CⅡTA的mRNA水平。结果pA629阳性ECV304细胞株与对照组比较,HLA-DR、-DP抗原表达分别降低了89.21%及92.31%;同时CⅡTA的mRNA含量降低(P<0.05)。结论CⅡTA的M1-RNA(M1-629-GS)降低了自身mRNA含量,从而阻止其调控的MHCⅡ类分子的表达。Objective To investigate the inhibitory effect of anfi-C Ⅱ TA M1-RNA on MHC Ⅱ expression. Methods M1-RNA with guide sequences （GS） recognized C Ⅱ TA at 452 and 629 site（M1-452-GS, M1- 629-GS, respectively） and C Ⅱ TA target RNA（114-800） were constructed, then cloned into the pUC19 and pGEM-7zf（ ＋ ） vector respectively. The recombinant M1-RNA and their target RNA were incubated in cell-free conditions. It showed that M1-629-GS could exclusively cleave target RNA. It was then cloned into the psNAV vector for intracellular analysis （ psNAV-M1-629-GS, pA629）. Stable transfeetants of ECV304 cell with pA629 were analyzed for classical MHC Ⅱ （HLA-DR, -DP, -DQ） expression by flow eytometry. The level of C Ⅱ TA mRNA was measured by RT-PCR. Results The expression of HLA-DR, -DP on pA629 positive ECV304 decreased 89.21% and 92.31% respectively . So did the mRNA amounts of C Ⅱ TA （ P 〈 0.05）. Conclusion M1-629-GS deduced C Ⅱ TA mRNA amounts. So classical MHC Ⅱ expression regulated by C Ⅱ TA was inhibited.

关 键 词：MHC Ⅱ类分子转录激活因子(CⅡTA) M1-RNA 移植免疫 

分 类 号：R392[医药卫生—免疫学]