鸟苷酸交换因子ARHGEF 10功能鉴定及其与肿瘤恶性表型间的关系研究

doi:10.11904/j.issn.1002-3070.2017.06.001

实用肿瘤学杂志 ›› 2017, Vol. 31 ›› Issue (6): 481-488.doi: 10.11904/j.issn.1002-3070.2017.06.001

• 基础研究 • 下一篇

鸟苷酸交换因子ARHGEF 10功能鉴定及其与肿瘤恶性表型间的关系研究

唐秋琳,毕锋

四川大学华西医院肿瘤分子靶向治疗研究室及肿瘤中心(成都 610041)

收稿日期:2017-06-26 出版日期:2017-12-28 发布日期:2018-01-02
通讯作者: 唐秋琳,E-mail:tangqiulin1981@163.com
作者简介:唐秋琳,女,(1981),硕士,实验师,从事肿瘤细胞生物学的研究。
基金资助:
国家自然科学基金面上项目(81572731);国家重点研发计划重点专项项目(2016YFC1303203)

Identification of guanine nucleotide exchange factor ARHGEF 10 and its relationship with tumor malignant phenotype

TANG Qiulin,BI Feng

Department of Laboratory Molecular Targeted Therapy of Tumor and Medical Oncology,West China Hospital,Sichuan University,Chengdu 610041,China

Received:2017-06-26 Online:2017-12-28 Published:2018-01-02

摘要/Abstract

摘要： 目的 Dbl家族鸟苷交换因子(GEFs)是Rho家族蛋白发生恶性转化的主要调控单位,它通过使Rho蛋白从无活性的GDP形式转换为GTP形式的Rho蛋白而发挥作用。本研究对一个GEF分子-ARHGEF 10进行了结构和功能上的初步探究,对该分子在肿瘤发展过程中扮演的角色进行讨论。方法 Realtime PCR对ARHGEF 10在人体42种正常组织中的表达情况进行了测定;GST-pulldown技术对ARHGEF 10的体内GEF活性进行了检测;双荧光素酶报告基因检测技术对下游小分子进行转录因子活性检测;应用免疫荧光双染标记法完成了高表达ARHGEF 10对正常细胞骨架形态的影响;在细胞表型实验中分别使用CCK8法、Transwell法及软琼脂克隆形成实验检测了高表达ARHGEF 10对细胞增殖侵袭迁移及体外成瘤能力的影响。结果生物信息学分析结果显示ARHGEF10分子量为156kD,具有典型的Dbl家族分子结构域,在肺组织中表达量最高,能够促使正常成纤维细胞中的应力纤维(Stress fiber)增多,同时促进细胞增殖、侵袭及克隆形成,体外转录因子活性检测发现该基因可能与JAK/STAT通路有关。结论 ARHGEF 10是一个典型的鸟苷交换因子家族分子,能激活Rho家族分子RhoA,具有明显的癌基因特征。

关键词: 鸟苷酸交换因子, ARHGEF 10, Dbl家族, Rho家族, 癌基因

Abstract: Objective The guanine exchange factors(GEFs)of Dbl family is a major regulatory unit for the malignant transformation of Rho family proteins.It plays a role by converting Rho protein from inactive GDP form to GTP form of Rho protein.In this paper,we discuss the structure and function of a GEF molecule-ARHGEF 10,and discuss its role in the process of tumor development.Methods The expression of ARHGEF 10 in 42 normal tissues was measured by Real-Time PCR.GST-pulldown technique was used to detect the GEF activity of ARHGEF 10 in vivo.The transcription factor activity of downstream small molecules was detected by dual-luciferase report gene assay.The high expressive effect of ARHGEF 10 on normal cytoskeleton morphology was performed by dual immunofluorescence staining labeling method.High expressive effects of ARHGEF 10 on cell proliferation,invasion and tumorigenic ability in vitro were examined using CCK8,Transwell and soft agar clony formation assays.Results ARHGEF 10 has a typical GEFs structure,which binds to RhoA in vitro and promotes the proliferation and invasion of NIH3T3 cells,and has significant ability to clone in vitro.Conclusion ARHGEF 10 is a typical family molecule of guanosine exchange factor that activates RhoA of Rho family,which has obvious oncogene characteristics.

Key words: Guanine nucleotide exchange factors, ARHGEF 10, Dbl, Rho GTPases, Oncogene

中图分类号:

R730.2

唐秋琳,毕锋. 鸟苷酸交换因子ARHGEF 10功能鉴定及其与肿瘤恶性表型间的关系研究[J]. 实用肿瘤学杂志, 2017, 31(6): 481-488.

TANG Qiulin,BI Feng. Identification of guanine nucleotide exchange factor ARHGEF 10 and its relationship with tumor malignant phenotype[J]. PRACTICAL ONCOLOGY JOURNAL, 2017, 31(6): 481-488.

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鸟苷酸交换因子ARHGEF 10功能鉴定及其与肿瘤恶性表型间的关系研究

Identification of guanine nucleotide exchange factor ARHGEF 10 and its relationship with tumor malignant phenotype

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