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MiR-300 regulates cellular radiosensitivity through targeting p53 and apaf1 in human lung cancer cells.

ABSTRACT: microRNAs (miRNAs) play a crucial role in mediation of the cellular sensitivity to ionizing radiation (IR). Previous studies revealed that miR-300 was involved in the cellular response to IR or chemotherapy drug. However, whether miR-300 could regulate the DNA damage responses induced by extrinsic genotoxic stress in human lung cancer and the underlying mechanism remain unknown. In this study, the expression of miR-300 was examined in lung cancer cells treated with IR, and the effects of miR-300 on DNA damage repair, cell cycle arrest, apoptosis and senescence induced by IR were investigated. It was found that IR induced upregulation of endogenous miR-300, and ectopic expression of miR-300 by transfected with miR-300 mimics not only greatly enhanced the cellular DNA damage repair ability but also substantially abrogated the G2 cell cycle arrest and apoptosis induced by IR. Bioinformatic analysis predicted that p53 and apaf1 were potential targets of miR-300, and the luciferase reporter assay showed that miR-300 significantly suppressed the luciferase activity through binding to the 3'-UTR of p53 or apaf1 mRNA. In addition, overexpression of miR-300 significantly reduced p53/apaf1 and/or IR-induced p53/apaf1 protein expression levels. Flow cytomertry analysis and colony formation assay showed that miR-300 desensitized lung cancer cells to IR by suppressing p53-dependent G2 cell cycle arrest, apoptosis and senescence. These data demonstrate that miR-300 regulates the cellular sensitivity to IR through targeting p53 and apaf1 in lung cancer cells.

SUBMITTER: He J

PROVIDER: S-EPMC5638365 | biostudies-literature | 2017 Oct

REPOSITORIES: biostudies-literature

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miR-300 regulates cellular radiosensitivity through targeting p53 and apaf1 in human lung cancer cells.

He Jinpeng J Feng Xiu X Hua Junrui J Wei Li L Lu Zhiwei Z Wei Wenjun W Cai Hui H Wang Bing B Shi Wengui W Ding Nan N Li He H Zhang Yanan Y Wang Jufang J

Cell cycle (Georgetown, Tex.) 20170912 20

microRNAs (miRNAs) play a crucial role in mediation of the cellular sensitivity to ionizing radiation (IR). Previous studies revealed that miR-300 was involved in the cellular response to IR or chemotherapy drug. However, whether miR-300 could regulate the DNA damage responses induced by extrinsic genotoxic stress in human lung cancer and the underlying mechanism remain unknown. In this study, the expression of miR-300 was examined in lung cancer cells treated with IR, and the effects of miR-300 ...[more]

PMID: 28895780

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Project description:ObjectiveTo determine the function of miR-125a-5p in laryngeal squamous cell carcinoma (LSCC), its correlation with radiation sensitivity, and the underlying regulatory mechanism.Materials and methodsWe conducted the analysis on the correlation between miR-125a-5p and head and neck squamous cell carcinoma (HNSCC) using data obtained from The Cancer Genome Atlas (TCGA). The putative gene targeted by miR-125a-5p has been identified as HK2, while the expression levels of miR-125a-5p and HK2 were measured in laryngeal cancer tissues and cells using RT-PCR. MiR-125a-5p and HK2 were introduced into the lentiviral vector and the vector was used to transfect AMC-HN-8 cells. The roles of miR-125a-5p and HK2 in LSCC and on radiosensitivity were determined by evaluating cell growth, examining colony formation, analyzing flow cytometry, and utilizing the single hit multi-target model. Western blotting was used to measure H2AX and rH2AX levels in the DNA damage response (DDR) pathway. The validation of the interaction between miR-125a-5p and HK2 was conducted through the dual-luciferase assay. To further confirm the association between miR-125a-5p and HK2, as well as its influence on radiosensitivity, rescue experiments were performed.ResultsThe expression of miR-125a-5p is downregulated in LSCC, while upregulating its expression could suppress cell growth, induce apoptosis, and enhance radiosensitivity. Additionally, HK2 exhibited high expression in LSCC and the biological function was opposite to miR-125a-5p. Western blotting analysis revealed that miR-125a-5p increased rH2AX levels and decreased H2AX levels, conversely, HK2 had the opposite effect on miR-125a-5p. These findings suggested that HK2 may serve as the target gene of miR-125a-5p. The double luciferase assay confirmed the binding of HK2 to miR-125a-5p, and rescue trials confirmed the role of miR-125a-5p in regulating the effects and radiation sensitivity of LSCC by targeting HK2 via the DDR pathway.ConclusionBy targeting HK2 and impacting the DDR pathway, miR-125a-5p has been found to inhibit cellular proliferation, enhance apoptosis, and heighten radiosensitivity in LSCC.

Dataset Information

MiR-300 regulates cellular radiosensitivity through targeting p53 and apaf1 in human lung cancer cells.

Publications

miR-300 regulates cellular radiosensitivity through targeting p53 and apaf1 in human lung cancer cells.

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