miR-204对TFAM的靶向调控作用及其对乳腺癌细胞生长与增殖的影响
| 作者: |
1肖顺崇,
1罗汉传,
1覃俊仕
1 广西医科大学第八附属医院/贵港市人民医院 甲状腺乳腺外科，广西 贵港 537100 |
| 通讯: |
罗汉传
Email: luohanchuan@126.com |
| DOI: | 10.3978/.10.3978/j.issn.1005-6947.2016.11.016 |
摘要
目的：探讨乳腺癌细胞中miR-204对线粒体转录因子A（TFAM）的靶向调控作用及其与细胞生长、增殖的关系。方法：将人乳腺癌MDA-MB-231细胞分别转染miR-204模拟物或miR-204抑制物，用real-time PCR和Western blot分别检测miR-204与TFAM蛋白的表达；构建荧光酶报告基因质粒（mut-TFAM/wt-TFAM），将其与miR-204模拟物或miR-204抑制物共转染MDA-MB-231细胞后检测荧光酶活性变化；构建pcDNA3.1/TFAM质粒，将其单独或与miR-204模拟物共转染MDA-MB-231细胞后检测TFAM蛋白表达，并用MTT法和BrdU法检测细胞生长与增殖情况。结果：MDA-MB-231细胞转染miR-204模拟物后miR-204的表达明显升高，而TFAM蛋白表达明显降低，转染miR-204抑制物后则呈反向变化（均P<0.05）。wt-TFAM与miR-204模拟物共转染时荧光酶活性明显下降，与miR-204抑制物共转染时荧光酶活性明显升高（均P<0.05）。转染pcDNA3.1/TFAM后，MDA-MB-231细胞的TFAM mRNA及蛋白表达量明显上调，细胞生长与增殖能力明显升高（均P<0.05）；miR-204模拟物后，MDA-MB-231细胞在TFAM表达降低的同时，细胞生长与增殖能力明显降低，而与pcDNA3.1/TFAM共转染后其上述作用均被部分抵消（均P<0.05）。结论：miR-204能靶向抑制乳腺癌细胞TFAM的表达，从而抑制乳腺癌细胞的生长与增殖。
关键词：
乳腺肿瘤
微RNAs
高迁移率族蛋白质类
细胞增殖
Targeted regulation of miR-204 on TFAM and their influence on growth and proliferation in breast cancer cells
CorrespondingAuthor:LUO Hanchuan Email: luohanchuan@126.com
Abstract
Objective: To investigate the targeted regulation of miR-204 on mitochondrial transcription factor A (TFAM) in breast cancer cells and their relations with cell growth and proliferation. Methods: Human breast cancer MDA-MB-231 cells were transfected with miR-204 mimics or inhibitors, and then, the miR-204 and TFAM protein expressions were determined by real-time PCR and Western blot, respectively. The luciferase reporter plasmids (mut-TFAM/wt-TFAM) were constructed and co-transfected with miR-204 mimics or inhibitors into MDA-MB-231 cells, and then, changes in luciferase activities were detected. The pcDNA3.1/TFAM plasmids were constructed and transfected alone or co-transfected with miR-204 mimics into MDA-MB-231 cells, and then, the TFAM protein expressions were measured, and cell growth and proliferation were analyzed by TTC and BrdU assay. Results: The miR-204 mRNA expression was significantly increased, and TFAM protein expression was significantly decreased in MDA-MB-231 cells after transfection with miR-204 mimics, while, opposite directional changes were found after transfection with miR-204 inhibitors (all P<0.05). The luciferase activity was significantly decreased after transfection with miR-204 mimics, but was significantly increased after transfection with miR-204 inhibitors (both P<0.05). In MDA-MB-231 cells, both expressions of TFAM mRNA and protein were significantly up-regulated, and the growth and proliferation were significantly enhanced after transfection of pcDNA3.1/TFAM (all P<0.05), and the growth and proliferation were significantly impaired along with significant down-regulation of TFAM protein expression after transfection of miR-204 mimics, which were all partially abolished by co-transfection with pcDNA3.1/TFAM (all P<0.05). Conclusion: MiR-204 exerts targeted inhibition on TFAM expression in breast cancer cells, and thereby suppresses the growth and proliferation of breast cancer cells.
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