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MicroRNA-181a Functions as an Oncomir in Gastric Cancer by Targeting the Tumour Suppressor Gene ATM

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Pathology & Oncology Research

Abstract

Based on our previous experiments, this study is to further investigate the functional significance of miR-181a and its target gene in gastric cancer. Expression of miR-181a was detected by qRT-PCR in three normal gastric tissues and three human gastric cancer cell lines (SGC-7901, MGC-803, and BGC-823 cells). After transfection with miR-181a inhibitor, proliferation, apoptosis, migration, and invasion of the SGC-7901 cells were evaluated. Ataxia-telangiectasia mutation (ATM) was predicted as a target gene of miR-181a with bioinformatics analysis, and was verified by lucifersae reporter assay. Expression of ATM protein in HEK293T cells and tissues was measured by Western Blot. Expression of ATM mRNA in HEK293T cells was measured by RT-PCR. Compared with three non-tumour tissues, the expression of miR-181a in three gastric cancer cells was significantly increased by 26.68, 14.83 and 14.96 folds; Compared with Negative Control(NC) and blank groups, transfection of miR-181a inhibitor led to inhibition of SGC7901 cell proliferation, invasion, and migration as well as promotion of apoptosis. A luciferase reporter assay demonstrated that ATM was a direct target of miR-181a, miR-181a mimics transfection down regulated ATM mRNA and protein expression. There was inverse correlation between miR-181a and ATM protein expression in gastric cancer and normal gastric tissues. Our study demonstrates that over-expression of miR-181a might be involved in development of gastric cancer by promoting proliferation and inhibiting apoptosis probably through directly targeting ATM. miR-181a modulation may be a potential strategy for the development of miRNA-based therapy of gastric cancer.

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Abbreviations

miRNA:

microRNA

BCL-2:

B cell leukemia-2

PCR:

Polymerase chain reaction

RT-PCR:

Reverse transcriptional PCR

CLL:

Chronic lymphatic leukemia

RISC:

RNA-induced silencing complex

PBS:

Phosphate balanced solution

NC:

Negative control

ATM:

Ataxia-telangiectasia mutated gene

rpm:

Revolutions per minute

bp:

Base pair

OD:

Optical density

ddH2O:

Double distilled water

cDNA:

Complementary deoxyribonucleic acid

pre-miRNA:

Precursor of miRNA

pri-miRNA:

Precursor of pre- miRNA

3′UTR:

3′-untranslated region

DEPC:

Dithylpyrocarbonate

GAPDH:

Glyceraldehyde-3-phosphate dehydrogenase

LNA:

Locked nucleic acid

DMSO:

Dimethyl sulphoxide

PLB:

Passive lysis buffer

BCA:

Bicinchoninic acid

SDS:

Sodium dodecyl sulfate

SNP:

Single nucleotide polymorphism

TBST:

Triethanolamine buffere saline solution with Tween-20

EDTA:

Ethylene diamine tetraacetic acid

MTS:

3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium

PI:

Propidium iodide

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Acknowledgements

This work was partly supported by the grant from Health bureau of Guangzhou City (NO.201102A212011) and by grant from the Natural Science Foundation of Guangdong Province (NO.10151006001000016)

Competing Interests Statement

The authors declare that they have no competing financial interests.

Author information

Authors and Affiliations

  1. Department of Gastroenterology, Shenzhen Futian Hospital of TCM, No. 6001 North Central Avenue of Futian, Shenzhen, China, 518034

    Xiangyang Zhang

  2. Department of Gastroenterology, Guangzhou Key Laboratory of Digestive Disease, Guangzhou First Municipal People’s Hospital, Guangzhou Medical University, No.1 Panfu Road, Guangzhou, China, 510180

    Yuqiang Nie, Jie Cao & Yanlei Du

  3. Department of Gastroenterology, The Fourth People’s Hospital of Shenzhen City (Futian Hospital), No. 3025 Middle Shennan Road, Shenzhen, China, 518033

    Xiaorong Li, Guifu Wu & Qun Huang

  4. Department of Gastroenterology, People’s Hospital Of New District Longhua Shenzhen, Jianshe East Road of Longhua New City, Shenzheng, China, 518019

    Junda Li & Ruoyu Deng

  5. Guangzhou Chest Hospital, Guangzhou, China, 510180

    Dongshen Huang

  6. Second People’s Hospital of Lanzhou City, Lanzhou, China, 730046

    Baozhi Chen

  7. Jiuquan City Health School, Lanzhou, China, 735000

    Shang Li

  8. Lanzhou Xigu People’s Hospital, Lanzhou, China, 730060

    Baojun Wei

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  1. Xiangyang Zhang
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  2. Yuqiang Nie
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  3. Xiaorong Li
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Correspondence to Yuqiang Nie.

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Zhang, X., Nie, Y., Li, X. et al. MicroRNA-181a Functions as an Oncomir in Gastric Cancer by Targeting the Tumour Suppressor Gene ATM. Pathol. Oncol. Res. 20, 381–389 (2014). https://doi.org/10.1007/s12253-013-9707-0

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  • DOI: https://doi.org/10.1007/s12253-013-9707-0

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