HPV阳性宫颈癌细胞中NFX1-123与HPV16 E6的细胞定位关系及其可能作用机制  

Relationship between NFX1-123 and HPV16 E6 in HPV-positive cervical cancer cells and its possible mechanism

作　　者：李娇生 郭一帆[2,3] 于浩天 LI Jiaosheng;GUO Yifan;YU Haotian(The Second School of Clinical Medicine,Southern Medical University,Guangdong Guangzhou 510168,China;Department of Obstetricsand Gynecology,Hainan Hospital of PLA General Hospital,Hainan Sanya 572013,China;Reproductive Medicine Center,Hainan Hospital of PLA General Hospital,Hainan Sanya 572013,China)

机构地区：[1]南方医科大学第二临床医学院,广东广州510168 [2]解放军总医院海南医院妇产科,海南三亚572013 [3]解放军总医院海南医院生殖医学中心,海南三亚572013

出　　处：《现代肿瘤医学》2023年第16期2972-2976,共5页Journal of Modern Oncology

基　　金：海南省自然科学基金面上项目(编号:820MS127)。

摘　　要：目的:构建并验证NFX1-123基因真核表达质粒,初探其在HPV阳性宫颈癌细胞中与HPV16 E6的细胞定位关系及可能作用机制。方法:获取NFX1-123基因片段,插入真核表达载体,构建并鉴定重组表达质粒。通过免疫共沉淀和免疫荧光方法对NFX1-123和HPV16 E6的相互作用与细胞定位关系进行研究。结果:酶切鉴定和测序确认了重组表达质粒nHA-NFX1-123/pRK5构建成功。NFX1-123蛋白可在HEK-293T细胞中正常表达且主要表达在细胞质中。通过免疫共沉淀法验证了NFX1-123和HPV16 E6蛋白结合。免疫荧光观察到与HPV16 E6共表达时,NFX1-123从细胞质迁移至细胞核并与细胞核中的HPV16 E6发生共定位。结论:成功构建nHA-NFX1-123/pRK5重组质粒。NFX1-123蛋白可与HPV16 E6蛋白结合,并受HPV16 E6影响从细胞质转移至细胞核,补充解释了为何胞质中的NFX1-123可与胞核中的HPV16 E6结合并通过多途径促进HR-HPV感染的宫颈上皮向宫颈癌发生。Objective:To construct and verify the eukaryotic expression plasmid of NFX1-123 gene,and to explore its cell localization relationship with HPV16 E6 in HPV positive cervical cancer cells and its possible mechanism.Methods:NFX1-123 gene fragment was obtained by PCR and inserted into the nHA/pRK5 vector.Restriction enzyme digestion and sequencing were used to identified the NFX1-123 recombinant plasmid.Western blot was used to confirm the overexpression of NFX1-123.Co-immunoprecipitation was used to certify its interaction with HPV16 E6.Results:The result of enzyme digestion and sequencing showed that NFX1-123 was successfully cloned into nHA/pRK5.Western blot showed that HA-NFX1-123 protein could be normally expressed in HEK-293T cells.The immunofluorescence revealed that NFX1-123 was mainly localized in the cytoplasm.The interaction between NFX1-123 and HPV16 E6 was demonstrated by co-immunoprecipitation.When HPV16 E6 was co-expressed,NFX1-123 underwent a change in localization from the cytoplasm to the nucleus and co-localized with HPV16 E6 in cell nucleus.Conclusion:Eukaryotic nHA-NFX1-123/pRK5 was successfully constructed.The interaction between NFX1-123 and HPV16 E6 was confirmed.HPV16 E6 could affect the subcellular localization of NFX1-123,shifting it from the cytoplasm to the nucleus.Supplementary explanation was provided on why NFX1-123 in cytoplasm can bind to HPV16 E6 in the nucleus and promote HR-HPV infected cervical epithelium to cervical cancer through various pathways.

关 键 词：NFX1-123 宫颈癌 高危型人乳头瘤病毒 E6 

分 类 号：R737.33[医药卫生—肿瘤]