微小 RNA-126-5p 通过直接靶向 CNOT7 抑制乳腺癌细胞的迁移。
MicroRNA-126-5p Inhibits the Migration of Breast Cancer Cells by Directly Targeting CNOT7.
机构信息
Department of Breast Surgery Ward, Jingjiang People's Hospital, Jingjiang, China.
Department of Blood Hernia Minimally Invasive Surgery, XuZhou Central Hospital, Xuzhou, China.
出版信息
Technol Cancer Res Treat. 2020 Jan-Dec;19:1533033820977545. doi: 10.1177/1533033820977545.
BACKGROUND
To assess the effect of microRNA-126-5p (miR-126-5p) on the migration of the breast cancer MCF7 cell line.
METHODS
GSE143564 was downloaded from the Gene Expression Omnibus (GEO; http://www.ncbi.nlm.nih.gov/geo) to identify the differentially expressed miRNAs between breast cancer and adjacent tissues. Quantitative reverse transcription PCR (RT-qPCR) was used to assess miR-126-5p levels in the normal 184A1 breast cell line and the breast cancer MCF7 cell line. The MCF7 cell line was then transfected with miR-126-5p mimics or corresponding negative control (NC-mimic). The proliferation and migration abilities of the MCF7 cell line were measured by methyl thiazolyl tetrazolium (MTT), Transwell and scratch healing assays. CCR4-NOT transcription complex and subunit 7 (CNOT7) expression levels in the NC-mimic and miR-126-5p mimic groups were measured by Western blot analysis. Bioinformatic analysis and a dual-luciferase reporter assay were performed to identify the miR-126-5p target gene.
RESULTS
One hundred forty-eight differentially expressed miRNAs (downregulated = 55, upregulated = 93) were identified. MiR-126-5p expression in the MCF7 cell line was significantly downregulated relative to that of 184A1 cell line (P < 0.05). Compared with that observed in the control and NC-mimic groups, cell proliferation in the miR-126-5p mimic group was significantly decreased at 48 and 72 h posttransfection (P < 0.05). In addition, the scratch healing rate and number of membrane-piercing cells in the miR-126-5p overexpression group were lower than those detected in the control and NC groups (P < 0.05). Furthermore, miR-126-5p could reduce the luciferase activity for the wild-type CNOT7 gene 3'-untranslated region (UTR) reporter (P < 0.05) but had no effect on the mutant 3'UTR reporter (P > 0.05). Compared with that observed in the NC and control groups, the levels of CNOT7 in the miR-126-5p overexpression group decreased (P < 0.05).
CONCLUSION
Upregulation of miR-126-5p can inhibit the migration of the breast cancer MCF7 cell line, which may involve its direct targeting of the 3'UTR of CNOT7.
背景
评估微小 RNA-126-5p（miR-126-5p）对乳腺癌 MCF7 细胞系迁移的影响。
方法
从基因表达综合数据库（GEO；http://www.ncbi.nlm.nih.gov/geo）下载 GSE143564，以鉴定乳腺癌与相邻组织之间差异表达的 miRNAs。采用定量逆转录 PCR（RT-qPCR）检测正常 184A1 乳腺细胞系和乳腺癌 MCF7 细胞系中 miR-126-5p 的水平。然后，用 miR-126-5p 模拟物或相应的阴性对照（NC-模拟物）转染 MCF7 细胞系。通过甲基噻唑基四唑（MTT）、Transwell 和划痕愈合测定法测量 MCF7 细胞系的增殖和迁移能力。通过 Western blot 分析测量 NC-模拟物和 miR-126-5p 模拟物组中 CCR4-NOT 转录复合物和亚基 7（CNOT7）的表达水平。通过生物信息学分析和双荧光素酶报告基因检测进行 miR-126-5p 靶基因鉴定。
结果
鉴定出 148 个差异表达的 miRNAs（下调=55，上调=93）。与 184A1 细胞系相比，MCF7 细胞系中 miR-126-5p 的表达显著下调（P<0.05）。与对照组和 NC-模拟物组相比，miR-126-5p 模拟物组在转染后 48 和 72 小时时细胞增殖明显减少（P<0.05）。此外，miR-126-5p 过表达组的划痕愈合率和穿膜细胞数均低于对照组和 NC 组（P<0.05）。此外，miR-126-5p 可降低野生型 CNOT7 基因 3'UTR 报告基因的荧光素酶活性（P<0.05），但对突变型 3'UTR 报告基因无影响（P>0.05）。与 NC 和对照组相比，miR-126-5p 过表达组中 CNOT7 的水平降低（P<0.05）。
结论
上调 miR-126-5p 可抑制乳腺癌 MCF7 细胞系的迁移，这可能涉及 miR-126-5p 对 CNOT7 的 3'UTR 的直接靶向作用。
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