Caspase-3融合蛋白基因的构建及其对HeLa细胞凋亡的诱导作用

目的：观察重构型caspase-3及其N-端融合蛋白的表达对HeLa细胞的凋亡诱导作用。方法：用反转录PCR获取人caspase-3基因，用重组PCR方法构建人重构型caspase-3及其融合蛋白基因，将基因克隆入表达载体pcDNA3，脂质体法转染HeLa细胞，通过细胞计数、电镜观察、MTT法等检测被转染细胞的生长及其凋亡状况。结果：成功获得了caspase-3基因，构建了重构型caspase-3基因和重构型caspase-3与绿脓杆菌外毒素（PE）转膜结构域部分肽段的融合蛋白基因及其表达载体，与对照组相比，在转染后1-4d内，两种基因的表达均导致HeLa细胞存活数减少，大量细胞发生凋亡，且二活性相当。结论：重构型caspase-3及其融合蛋白可以诱导转染的HeLa细胞发生凋亡。

Construction and effect of caspase-3 fusion proteins and their apoptosis induction in HeLa cells

AIM To investigate the induction of HeLa cells to apoptosis by the expression of reversed caspase 3 and its N terminal fusion protein. METHODS RT PCR and recombi nant PCR were used to amplify human caspase 3 gene and construct reversed caspase 3 and its fusion protein genes. Following cloning into expression vector pcDNA3 and transfecting HeLa cells, we evaluated the growth condition and apoptosis of these cells by cell counting, electron microscope photographs, as well as MTT assays. RESULTS We successfully obtained human caspase 3 gene, the genes and expression vectors of reversed caspase 3 and its fusion protein with part of the translocation domain of Pseudomonas exotoxin A (PE). Compared with the control group, the expression of both genes caused in a similar degree the decrease of living cell numbers and massive occurrence of apoptosis. CONCLUSION Both reversed caspase 3 and its fusion protein can effectively induce HeLa cells to apoptosis.