热休克蛋白70对突变型p53稳定性的影响

Effect of HSP70on Stability of Mutant p53Protein

目的:探讨热休克蛋白70(heatshockprotein70HSP70)对突变型p53(mutanttypep53mtp53)稳定性的影响。方法将反义HSP70真核表达重组质粒pXAHSP70脂质体介导转染至MCF7/Adr人乳腺癌细胞,PCR确定转染细胞中外源DNA存在,Westernblot检测HSP70蛋白的抑制程度,p53稳定性实验分析反义HSP70RNA对mtp53半衰期的改变。结果:转染重组质粒的MCF7/Adr细胞中HSP70蛋白表达下降42%,其胞内mtp53含量下降,半衰期缩短了6h,稳定性下降。结论:反义HSP70RNA能够明显地降低mtp53稳定性,提示肿瘤细胞中HSP70高表达可增强mtp53的稳定性。

Objective:The current study was designed to investigate the effect of heat shock pro tein 70(HSP70)on stability of mutant p53protein.Methods :Retroviral recombinant expressing antisense HSP70RNA was constructed and transfected into MCF7/Adr breast ca ncer cells.The existence of foreign DNA was identified by PCR method and t he HSP70protein level was determined by Western blot analysis.The half-life of m utant p53protein(mtp53)was measured by p53stability assay.Results:The stable expressing strain(MAp70)from transfected cells was obtained through G418selection.The foreign DNA in transfectant cells were confirmed by PCR,and the repre ssion rate of HSP70protein was 42%.T he half-life of mutant p53in MAp70cell was 12hours,significantly lower th an that of the control cells.Conclusion:Antisense HSP70RNA can decrease the HSP70protein level and significant ly increase instability of mutant p53protein in MCF7/Adr breast cancer cell.