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hTERT上调FOXM1乙酰化影响胃癌细胞侵袭能力
hTERT promotes FOXM1 acetylation and subsequent invasion of gastric cancer cells
- 第三军医大学学报 2017年39卷第23期 页码：2243-2249
- 作者机构：
400037 重庆，第三军医大学新桥医院消化内科
- 作者简介：
[柏健鹰，E-mail:baijianying@tmmu.edu.cn BAI Jianying, baijianying@tmmu.edu.cn
- 基金信息：重庆市基础科学与前沿技术研究项目(CSTC2016jcyjA0570)
DOI：10.16016/j.1000-5404.201708132
中图分类号： R345;R394.3;R735.2收稿：2017-08-22，
修回：2017-09-18，
纸质出版：2017-12-15
- 稿件说明：
移动端阅览
廖国斌, 彭瑞琨, 节梦梦, 等. hTERT上调FOXM1乙酰化影响胃癌细胞侵袭能力[J]. 第三军医大学学报, 2017,39(23):2243-2249.
Guobin LIAO, Ruikun PENG, Mengmeng JIE, et al. hTERT promotes FOXM1 acetylation and subsequent invasion of gastric cancer cells[J]. Journal of Third Military Medical University, 2017, 39(23): 2243-2249.
廖国斌, 彭瑞琨, 节梦梦, 等. hTERT上调FOXM1乙酰化影响胃癌细胞侵袭能力[J]. 第三军医大学学报, 2017,39(23):2243-2249. DOI： 10.16016/j.1000-5404.201708132.
Guobin LIAO, Ruikun PENG, Mengmeng JIE, et al. hTERT promotes FOXM1 acetylation and subsequent invasion of gastric cancer cells[J]. Journal of Third Military Medical University, 2017, 39(23): 2243-2249. DOI： 10.16016/j.1000-5404.201708132.
摘要
目的
2
探讨人端粒酶逆转录酶(human telomerase reverse enzyme，hTERT)促进胃癌细胞侵袭的分子机制。
方法
2
构建稳定过表达hTERT的细胞株SGC-7901-hTERT，采用Western blot检测叉头盒蛋白(forkhead box M1，FOXM1)的表达，qRT-PCR和Western blot检测FOXM1下游侵袭相关基因基质金属蛋白酶-2(matrix metalloproteinase-2，MMP-2)、基质金属蛋白酶-9(MMP-9)的mRNA和蛋白表达；构建FOXM1的shRNA干扰质粒，在SGC-7901-hTERT细胞中转染该质粒，Transwell法检测细胞侵袭能力，qRT-PCR和Western blot检测MMP-2、MMP-9的mRNA和蛋白表达；构建FOXM1启动子活性报告质粒，在SGC-7901-hTERT细胞中用双荧光素酶报告实验检测FOXM1启动子活性，qRT-PCR检测FOXM1 mRNA表达；构建稳定过表达FOXM1的细胞株SGC-7901-HA-FOXM1，并在此细胞中转染hTERT过表达质粒，免疫共沉淀(co-immunoprecipitation，Co-IP)检测FOXM1乙酰化水平变化，免疫荧光实验检测hTERT与FOXM1的细胞定位。
结果
2
与对照组比较，过表达hTERT促进FOXM1及其下游分子MMP-2、MMP-9的蛋白表达，同时也促进肿瘤细胞侵袭(
P
<
0.01)，但不影响FOXM1的启动子活性及mRNA表达；与过表达hTERT组比较，干扰FOXM1可抑制MMP-2、MMP-9的mRNA和蛋白表达，同时也抑制肿瘤细胞侵袭(
P
<
01)；hTERT与FOXM1存在细胞共定位，且过表达hTERT可上调FOXM1乙酰化水平。
结论
2
hTERT通过上调FOXM1乙酰化促进胃癌细胞侵袭。
Abstract
Objective
2
To investigate the molecular mechanism of human telomerase reverse transcriptase (hTERT) in promoting the invasion of gastric cancer cells.
Methods
2
A cell line with stable expression of hTERT
SGC-7901-hTERT was constructed based on human gastric cancer SGC-7901 cells. Western blotting was used to detect the expression of forkhead box M1 (FOXM1)
and it combined with qRT-PCR to measure the expression of FOXM1 downstream molecules
matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) at protein and mRNA levels. The shRNA interference plasmid of FOXM1 was constructed and transfected into the SGC-7901-hTERT cells. Transwell chamber test was used to detect the invasive ability of the cells. qRT-PCR and Western blotting were used again for the expression of MMP-2 and MMP-9. The activity of FOXM1 promoter was detected by double luciferase reporter assay in SGC-7901-hTERT cells and corresponding control cells
and then the mRNA expression of FOXM1 was detected by qRT-PCR. After the construction of SGC-7901 FOXM1-HA cells with FOXM1-HA lentivirus
the cells were transfected with hTERT overexpression plasmid. The acetylation of FOXM1 was detected by co-immunoprecipitation (Co-IP). The immunofluorescence assay was used to detect the co-localization of hTERT and FOXM1.
Results
2
Compared with the control group
hTERT overexpression promoted the expression of FOXM1 protein
which facilitated the expression of FOXM1 target genes
MMP-2 and MMP-9
and also promoted tumor cell invasion (
P
<
01). However
the expression of FOXM1 mRNA and its promoter activity was not affected. Compared with the overexpressing hTERT group
interference of FOXM1 partially decreased the effect of hTERT-promoted FOXM1 target gene expression and reduced the hTERT-mediated invasion of SGC-7901 cells (
P
<
01). FOXM1 and hTERT were co-localized in the SGC-7901 cells
and overexpression of hTERT up-regulated the level of acetylation of FOXM1.
Conclusions
2
hTERT affects the invasion of gastric cancer cells by up-regulation of FOXM1 acetylation.
关键词
Keywords
references
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