精胺对Nup98基因的调控研究

Spermine Regulating the Nup 98 Expression in NIH3T3 Cells Through Enhancing Its Promoter Activity

利用基因芯片比较鸟氨酸脱羧酶抑制子基因敲除小鼠和正常小鼠肝组织的表达差异;高效液相色谱法测定鸟氨酸脱羧酶抑制子基因敲除小鼠和正常小鼠肝组织中的精胺含量;构建Nup98的启动子-虫荧光素酶报告基因质粒;脂质体Lipofectamine 2000TM转染小鼠细胞(NIH3T3),在含有精胺或正常培养条件下,测定虫荧光素酶活性。结果显示,Nup98基因在鸟氨酸脱羧酶抑制子基因敲除小鼠肝组织表达增强;鸟氨酸脱羧酶抑制子基因敲除小鼠肝组织中,精胺含量增高;成功构建Nup98的启动子-虫荧光素酶报告基因质粒pGL3-Nup98-P;精胺能在体外培养的条件下,增强虫荧光素酶的活性。这些结果证明精胺能激活Nup98基因的启动子活性,增强其在NIH3T3细胞中的表达。

The Nup98 expressions between Azin1 knock out mice and wild-type mice were analyzed by using Real-time PCR.The levels of spermine in Azin1 knock out mice and in wild-type mice were determined by HPLC method.Nup98 promoter luciferase reporter plasmid was constructed,then NIH3T3cells were transiently transfected with the same amount of Nup98 promoter luciferase reporter plasmid,and luciferase activity was detected with and without spermine.Results showed that Nup98 expression on mRNA level is significantly elevated in liver of Azin1 knock out mice.The level of spermine was increased in Azin1 knock out mice.Nup98 promoter luciferase reporter plasmid pGL3-Nup98-P,was constructed successfully.Spermine could enhance the activity of Nup98 promoter.These data demonstrated that spermine regulates the Nup98 expression in NIH3T3 cells through enhancing its promoter activity.