PLAGL2对SP-C基因表达的调控及其机制的初步研究

目的研究多形性腺瘤样因子2（pleiomorphic adenoma gene like -2, PLAGL2）对肺表面活性蛋白C基因（surfactant protein-C，SP-C）表达的影响及调控机制。方法通过定点诱变技术分别突变掉SP-C基因启动子上PLAGL2的结合位点以及PLAGL2的第6、7位锌指结构域并获得相应的突变体；利用细胞共转染和荧光素酶报告基因检测技术研究PLAGL2对SP-C基因启动子表达活性的影响及比较正常及突变体的PLAGL2或SP-C基因启动子之间相互作用的差异以明确PLAGL2对SP-C基因的调控及作用靶点。结果细胞共转染试验中，荧光素酶活性检测结果表明PIAGL2可明显增加SP-C基因的表达（P〈0．001），SP-C基因表达活性的增高与PLAGL2的作用浓度呈正相关；当定点诱变掉PLA-GL2的第6或7位锌指结构域后，PLAGL2的突变体质粒均不能增高SP-C基因的表达活性，与对照组相比较，差异具有显著性（P〈0．001）；同样，当定点诱变掉PIAGL2在SP-C基因启动子上的结合位点后，PLAGL2也不能增高SP-C基因启动子突变体质粒的表达活性，与对照组相比较，两者具有明显差异（P〈0．01）。结论在肺Ⅱ型细胞中，PLAGL2通过其第6和7位锌指结构域结合于SP-C基因启动子上的PLAGL2结合位点序列来促进SP-C基因的表达。

Regulation mechanism of pleiomorphic adenoma gene like-2 modulating surfactant protein-C gene expression in the lung cells

Objective： To investigate the regulation mechanism of PLAGL2 modulating SP- C gene expression. Methods： Site- directed mutagenesis was performed to get the SP - C gene promoter mutant （ SP - CP - mComclu ） and PLAGL2 mutants （ PLAGL2 - mF6, PLAGL2 - mFT） ; Co - transfaction study and luciferase activity measurement were employed to study the effect on sp - c gene expression exerting by PLAGL2 and to analysis the activity difference of sp - c gene expression made by wild type and mutants PLAGL2, so as to identify the DNA - binding ability and specifity of interaction between PLAGL2 and SP - C gene promoter. Results ： Luciferase re- suits showed that PLAGL2 greatly transactivated the SP -C gene promoter in co -transfaction study （ P 〈0. 001 ）, and the activation is PLAGL2 concentration dependent. PLAGL2 - mF6 or PLAGL2 - mF7 did not increase the regulation activity of SP - C gene promoter expression compared with control （ P 〈 0. 001 ） ; PLAGL2 also can not increase the regulation activity of SP - CP - mComclu promoter expression compared with control （ P 〈0. 01 ）. Conclusion： PLAGL2 could greatly promote the SP -C gene activity in the lung cells, it can bind to the core and cluster binding site on the SP - C gene promoter by it＇s zinc finger 6 and 7 to affect SP - C gene expression.