iNOS基因在胶质细胞中转录激活机制初探

Mechanism of Transcriptional Activation of iNOS in Glial Cell

目的:通过瞬时转染p38MAPK途径中上游激酶:组成激活型MAPK激酶3(MKK3b)和MAPK激酶6(MKK6b),进一步了解p38MAPK级联传导信号系统调节iNOS基因在胶质细胞中的转录激活机制。方法:MKK3b或MKK6b与接有荧光素酶(luciferase,Luc)的大鼠iNOS启动基因质粒(iNOS-Luc);cAMP反应元件(CRE-Luc)和核因子κB(NFκB-Luc)联合转染C6胶质细胞株。结果:MKK3b/MKK6b能引起iNOS-Luc的激活,并都能够被p38MAPK抑制剂SB203580所抑制。MKK可以诱导cAMP反应元件(CRE-Luc)介导的和核因子κB(NFκB-Luc)依赖的转录活性。显性抑制型(dominantnegative,dn)CRE结合蛋白(dnCREB)和CCAAT/增强子结合蛋白(C/EBP)都是p38MAPK的靶向作用目标。结论:转染这两种转录因子产生相反的影响:dnCRE增强了iNOS-Luc的表达,而dnC/EBP则引起抑制作用,对CRE-Luc也具有相同的影响。

Objective：To investigate the role of p38 MAPK in the transcriptional activation of the iNOS gene. Methods： Transient transfection with constitutively active upstream kinases in the pathway, i.e. MKK3b（E） and MKK6b（E）. Expression in C-6 glial cells of either MKK3b（E） or MKK6b（E） . Results： An induction of the activity of a cotransfected rat iNOS promoter-reporter （iNOS-Luc） gene and an enhancement of expression of iNOS mRNA, both of which were inhibitable by the p38 MAPK inhibitor SB203580. Conclusion： The MKK constructs induced cAMP response element-mediated （CRE-Luc） and nuclear factor KB-dependent （nuclear factor KB-Luc） transcriptional activities. Transfection with dominant negative （dn） forms of CRE-binding protein （CREB） and CCAAT/enhancerbinding protein （C/EBP）, the two CRE-binding transcription factors targeted by the p38 MAPK pathway, resulted in opposite effects; dnCREB enhanced and dnC/EBP inhibited iNOS-Luc parallel to their effects on CRE-Luc.