|本期目录/Table of Contents|

[1]金泽源,李咏梅.Fus依赖ac4C修饰调控促进宫颈癌细胞增殖[J].天津医科大学学报,2023,29(01):15-20.
　JIN Ze-yuan,LI Yong-mei.Fus promotes cell proliferation by ac4C-modification in cervical cancer cells[J].Journal of Tianjin Medical University,2023,29(01):15-20.
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《天津医科大学学报》[ISSN:1006-8147/CN:12-1259/R]

卷:
29卷
期数:
2023年01期
页码:
15-20
栏目:
肿瘤基础研究专题
出版日期:
2023-01-20

文章信息/Info

Title:
Fus promotes cell proliferation by ac4C-modification in cervical cancer cells
文章编号:
1006-8147(2023)01-0015-06
作者:
金泽源; 李咏梅
（天津医科大学基础医学院病原生物学系，天津 300070）
Author(s):
JIN Ze-yuan; LI Yong-mei
（Department of Pathogenic Biology，School of Basic Medical Sciences，Tianjin Medical University，Tianjin 300070，China）
关键词:
宫颈癌; ac4C; Fus; NAT10
Keywords:
cervical cancer; ac4C; Fus; NAT10
分类号:
R737.33
DOI:
-
文献标志码:
A
摘要:
目的：探讨Fus RNA 结合蛋白（Fus）影响宫颈癌细胞增殖的机制。方法：通过MTT实验观察Fus对宫颈癌细胞HeLa增殖的影响；采用RT-qPCR和Western印迹观察人N-乙酰转移酶10（NAT10）对Fus表达的影响；利用RNA免疫沉淀实验（RIP）检测Fus mRNA的富集情况；通过RNA稳定性实验观察NAT10对Fus mRNA稳定性的影响；基于生物信息学软件PACES预测Fus mRNA上的ac4C修饰位点，构建含有Fus mRNA上ac4C修饰位点的野生型和突变型的EGFP报告质粒，利用EGFP-reporter实验和乙酰化RIP（ac4C-RIP）确定Fus mRNA上发生ac4C修饰位点；最后通过挽救实验观察NAT10介导Fus对宫颈癌细胞增殖的影响。结果：Fus促进宫颈癌HeLa细胞增殖（tFus=14.06，tshR-Fus=5.74，均P<0.05）；Fus蛋白和mRNA的表达水平随NAT10表达增加而增加（tNAT10=10.71，tshR-NAT10-1=13.17，tshR-NAT10-2=7.65，均P<0.05）；RIP实验结果提示Fus mRNA与NAT10蛋白间存在相互作用（t=4.20，P<0.05），且NAT10过表达细胞中Fus mRNA具有较高ac4C修饰水平（t=30.62，P<0.05）；另外，相对于载体对照细胞，NAT10过表达细胞中Fus mRNA的稳定性增高（P<0.01）；发现Fus mRNA的925～939序列中的胞嘧啶为ac4C修饰位点；挽救实验结果显示NAT10介导Fus促进宫颈癌细胞的增殖（t=9.67、6.71，P<0.05）。结论：Fus mRNA被NAT10催化发生ac4C修饰，Fus表达水平被上调，继而促进宫颈癌细胞的增殖。
Abstract:
Objective： To explore the mechanism of Fus RNA binding protein（Fus） in the promotion of cervical cancer cell proliferation. Methods：The effects of Fus on cervical cancer cell proliferation were observed using MTT assays. Western blotting analysis and RT-qPCR were used to detect the influence of human N-acetyltransferase 10（NAT10） on expression levels of Fus. RNA immunoprecipitation assay （RIP） was detected the enrichment of Fus mRNA. RNA half-life assay was conducted to determine the effect of NAT10 on the stability of Fus mRNA. Based on the searching results of the possible ac4C-modification sites on Fus mRNA through PACES bioinformatics software，the EGFP reporter plasmids containing the possible wild-type or mutant ac4C-modification sites were constructed. EGFP-reporter assay and ac4C-RIP assay were used for the identification of ac4C modification sites on Fus mRNA. Lastly，the effect of NAT10 mediated Fus on the proliferation of cervical cancer cells was observed by rescue experiment. Results：Fus promoted cell proliferation of HeLa cells（tFus=14.06，tshR-Fus=5.74，all P<0.05）. The expression level of Fus protein and mRNA was increased with NAT10 expression（tNAT10=10.71，tshR-NAT10-1=13.17，tshR-NAT10-2=7.65，all P<0.05）. The results of RIP experiment indicated that there was an interaction between Fus mRNA and NAT10 protein（t=4.20，P<0.05），and Fus mRNA in NAT10 overexpression cells had a higher ac4C modification level（t=30.62，P<0.05）. In addition，the stability of Fus mRNA in NAT10 overexpression cells was higher than that in vector control cells（P<0.01） Furthermore，cytosine in 925-939 sequence of Fus mRNA was ac4C modified site. The rescue experiment results showed that NAT10 mediated Fus to promote the proliferation of cervical cancer cells（t=9.67，6.71，both P<0.05）. Conclusion：Fus mRNA can be ac4C modificated by NAT10，which resulting in the enhancement of Fus expression and the promotion of cell proliferation in cervical cancer cells.

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备注/Memo

备注/Memo:
作者简介:金泽源（1995-），男，硕士在读，研究方向：病原生物学； 通信作者：李咏梅，E-mail：liym@tmu.edu.cn。
更新日期/Last Update: 2023-02-01