目的 分析衰老相关蛋白prelamin A是否与MT-2A相互作用,以及MT-2A与衰老的关系.方法 采用酵母双杂交方法从人骨骼肌文库中筛选prelamin A的相互作用蛋白,酵母一对一回复性杂交和荧光共定位验证MT-2A与prelamin A是否相互作用.RT-PCR分析MT-2A在正常和早老细胞中的表达.结果 通过酵母双杂交方法从人骨骼肌文库中筛选到prelamin A的候选相互作用蛋白MT-2A,一对一回复性验证也证明两者能在酵母细胞中相互作用.构建含绿色荧光蛋白的MT-2A融合表达载体转染HEK-293细胞,激光共聚焦显微观察发现MT-2A能与带红色荧光蛋白的prelamin A蛋白在核膜共定位.研究还发现MT-2A的表达在早老细胞中显著下调(P＜0.01).结论 MT-2A作为一个新的prelamin A结合蛋白,可能在细胞早老过程中具有重要的作用.%Objective To analyze whether prelamin A interacts with MT-2A and the relationship of MT-2A with senescence. Methods Prelamin A binding partners were searched by a yeast two-hybrid screen using a human skeletal muscle cDNA library and validated the interaction of MT-2A with prelamin A by one to one reverse hybridization and co-localization experiment. MT-2A mRNA level was analyzed by RT-PCR. Results MT-2A was identified to interact with prelamin A by yeast two-hybrid screen and one to one reverse hybridization. MT-2A and prelamin A were co-located around nuclear envelope using laser confocal microscopy when co-transfected with pDsRed-prelamin A and pEGFP-MT-2A. RT-PCR results showed that the mRNA level of MT-2A in HEK-293prelamin A cells was significantly lower than that in HEK-293 cells (P <0. 01). Conclusions MT-2A, as a novel prelamin A binding partner, might play a role in premature senescence.
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