JWA基因的蛋白缺陷对苯并(a)芘所致HeLa细胞DNA损伤的影响

目的研究JWA基凶表达缺陷对苯并（a）芘诱导细胞DNA损伤与修复的影响肢可能机制。方法构建JWA基因反义RNA绿色荧光蛋白真核表达载体（pEGFP—C1-asJWA）,用该载体稳定转染人宫颈癌细胞（HeLa）获得JWA蛋白缺陷细胞株（asJWA-HeLa）,在含有S9的培养条件下以50μmol/L苯并（a）芘处理细胞3h,再恢复不同时间,用碱性单细胞凝胶电泳方法鉴定DNA损伤程度。结果asJWA-HeLa细胞JWA蛋白表达水平比对照下降了69％。在苯并（a）芘致DNA损伤模型中,asJWA-HeLa细胞的DNA损伤程度重于对照细胞,并且出现明显的DNA修复延迟现象。结论JWA作为一种新的环境应答基因,活跃地参与了举并（a）芘诱导的HeLa细胞DNA损伤与修复过程,并住其中起着积极的保护作用。

Expression deficiency of JWA enhanced DNA damage and delayed DNA repair in HeLa cells induced by benzo (a) pyrene exposure

OBJECTIVE: To investigate the role of JWA gene in benzo (a) pyene B (a) P] induced DNA damage and repair effects in HeLa cells. METHODS: The antisense JWA express vector (pEGFP-C1-asJWA) was constructed and stably transfected into HeLa cells. JWA deficient HeLa cells (asJWA-HeLa) was then screened and established. The general characteristics of asJWA-HeLa cells were investigated. DNA damage and repair cell culture model was conducted by treating the cells with 50 micromol/L B (a) P plus S9 for 3 hours and then the cells were maintained further 0, 1, 3, and 24 hours for DNA repairing. The damaged DNA was detected by single cell gel electrophoresis assay (comet assay). RESULTS: JWA deficient HeLa cells (with a 31% of JWA protein expression as compared with the control) were obtained successfully. Compared with the empty vector transfcted cells (C1-HeLa) and the untransfected HeLa cells, asJWA-HeLa cells were more sensitive to B (a) P exposure and with a delayed DNA repair process. CONCLUSION: The JWA determined might function as a potential effective environmental responsive gene and actively participate the process of B (a) P exposure associated with intracellular signal pathways of DNA damage and repair.