文章摘要
奚佩雯,胡 玥,张 旭,戴欣媛,石 靓,丁 强.RBM7在乳腺癌细胞BT474中的表达及其与P21的关系[J].南京医科大学学报,2020,(2):160~165
RBM7在乳腺癌细胞BT474中的表达及其与P21的关系
The expression of RBM7 and its relationship with P21 in breast cancer cell BT474
投稿时间:2019-06-30  
DOI:10.7655/NYDXBNS20200203
中文关键词: 乳腺癌  RNA结合蛋白7  P21  细胞周期
英文关键词: breast cancer  RNA binding protein 7  P21  cell cycle
基金项目:国家自然科学基金(81572595)
作者单位
奚佩雯 南京医科大学第一附属医院乳腺病中心 江苏 南京 210029 
胡 玥 南京医科大学第一附属医院乳腺病中心 江苏 南京 210029 
张 旭 南京医科大学第一附属医院乳腺病中心 江苏 南京 210029 
戴欣媛 南京医科大学第一附属医院乳腺病中心 江苏 南京 210029 
石 靓 南京医科大学第一附属医院乳腺病中心 江苏 南京 210029 
丁 强 南京医科大学第一附属医院乳腺病中心 江苏 南京 210029 
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中文摘要:
      目的:探索RNA结合蛋白7(RNA binding protein 7,RBM7)在人源性乳腺癌细胞BT474中的过表达及干扰后的表达改变,并研究其与P21之间的关系。方法:在人源性乳腺癌细胞BT474中分别转染RBM7过表达、干扰慢病毒(实验组)和相应对照慢病毒(对照组),从而构建稳定转染的细胞株,用qRT?PCR和Western blot实验分别验证转染后的RBM7的表达情况。用CCK?8实验、流式细胞周期实验分别评估RBM7表达改变后对乳腺癌细胞增殖能力和细胞周期分布的影响。通过qRT?PCR和Western blot实验观察BT474细胞中RBM7表达改变后对P21表达的影响。进一步通过RNA结合蛋白免疫共沉淀(RIP)实验来研究RBM7与P21之间的关系。结果:在乳腺癌细胞BT474中分别转染RBM7过表达、干扰慢病毒48 h后,通过荧光显微镜检测细胞的绿色荧光表达情况;经嘌呤霉素稳定筛选后,获得RBM7过表达及干扰的稳转细胞株。CCK?8和流式细胞周期实验显示,过表达RBM7后可促进乳腺癌细胞的增殖,而干扰RBM7后能抑制乳腺癌细胞的增殖。qRT?PCR和Western blot实验显示,过表达RBM7能下调P21的mRNA(P < 0.05)及蛋白的表达,干扰RBM7能上调P21的mRNA(P < 0.05)及蛋白的表达。RNA结合蛋白免疫共沉淀(RIP)实验显示,RBM7能直接结合P21的mRNA从而发挥其对P21的调节作用。结论:RBM7在人乳腺癌细胞BT474中通过结合P21的mRNA从而下调P21的表达。
英文摘要:
      Objective:This study aims to investigate the relationship between RNA binding protein 7(RBM7) and P21 via overexpressing and knocking down RBM7 in human breast cancer cell BT474. Methods:BT474 cells were respectively transfected with RBM7 overexpressing,knocking down lentivirus(experimental group)and corresponding control lentivirus(control group). The expression of RBM7 was verified after BT474 stably transfected with RBM7 letivirus by the qRT?PCR and Western blot assays. The effects of RBM7 overexpression or knock down on breast cancer proliferation were evaluated by using CCK?8 assays. At the same time,the flow cytometry assays were used to detect the cell cycle distribution of RBM7 overexpression or knock down. The qRT?PCR and Western blot assays were used to investigate the expression of P21 following the variation of RBM7. Furthermore,the relationship between RBM7 and P21 was studied by RNA binding protein immunoprecipitation(RIP) assays. Results:The green fluorescence was observed by fluorescence microscope when BT474 was transfected with RBM7 overexpression and knock down lentivirus after 48 h. After stably screening,the fluorescence expression of cells was significantly stronger. CCK?8 and the flow cytometry assays showed that overexpressing RBM7 could promote the proliferation of breast cancer cells,while knocking down RBM7 could inhibit the proliferation of breast cancer cells. It was observed that overexpression of RBM7 could downregulate mRNA(P < 0.05)and protein expression of P21,and knockdown of RBM7 upregulated mRNA(P < 0.05)and protein expression of P21 by qRT?PCR and Western blot. It was revealed that RBM7 could directly bind to mRNA of P21 by RNA?binding protein immunoprecipitation(RIP)assays. Conclusion:RBM7 can downregulate the expression of P21 by binding to its transcript in breast cancer cell BT474.
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