title: Long intergenic non-coding RNA induced by X-ray irradiation regulates DNA damage response signaling in the human bronchial epithelial BEAS-2B cell line.
abstract: Long non-coding RNAs (lncRNAs) have been regarded as the primary genetic regulators of several important biological processes. However, the biological functions of lncRNAs in radiation-induced lung damage remain largely unknown. The present study aimed to investigate the potential effects of lncRNAs on radiation-induced lung injury (RILI). Female C57BL/6 mice were exposed to 12 Gy single doses of total body irradiation (TBI). LncRNA microarray screening was conducted at 24 h post-irradiation (IR) to investigate the differentially-expressed lncRNAs during RILI. Following the subsequent bioinformatics analysis and reverse transcription-polymerase chain reaction (RT-PCR) validation, one of the verified differentially-expressed long intergenic radiation-responsive ncRNAs (LIRRs), LIRR1, was selected for further functional study. The normal human bronchial epithelial BEAS-2B cell line was used as the cell model. The recombinant eukaryotic expression vector for the lncRNA was designed, constructed and transfected using lipofectamine. RT-PCR, clonogenic and flow cytometry assays, immunofluorescence detection and western blot analysis were performed to reveal the role of the lncRNA in the radiosensitivity regulation of the RILI target cells. In lung tissues 24 h after 12 Gy TBI, six of the identified differentially-expressed LIRRs near the coding genes were validated using quantitative (q)PCR. The upregulation of two LIRRs was observed and confirmed using qPCR. LIRR1 was chosen for further functional study. Following the stable transfection of LIRR1, identified through G418 screening, increased radiosensitivity, evident cell cycle G1phase arrest and increased γ-H2AX foci formation were observed in the bronchial epithelial BEAS-2B cell line subsequent to IR. LIRR1 overexpression also led to decreased expression of the KU70, KU80 and RAD50 DNA repair proteins, marked activation of p53, decreased mouse double minute 2 homolog (MDM2) expression, and substantially induced p21 and suppressed cyclin-dependent kinase 2 in BEAS-2B following IR. Subsequent to the use of Pifithrin-α, a specific inhibitor of p53 activation, increased MDM2 expression was observed in the LIRR1-overexpressing cells, suggesting that LIRR1 could mediate the DNA damage response (DDR) signaling in a p53-dependent manner. The present study provides a novel mechanism for RILI, using the concept of lncRNAs.
authors.affiliation: Jiangsu Provincial Key Laboratory of Radiation Medicine and Protection, School of Radiation Medicine and Protection, Medical College of Soochow University, Suzhou, Jiangsu 215123, P.R. China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Jiao
authors.affiliation: Jiangsu Provincial Key Laboratory of Radiation Medicine and Protection, School of Radiation Medicine and Protection, Medical College of Soochow University, Suzhou, Jiangsu 215123, P.R. China.
authors.forename: Chang
authors.initials: C
authors.lastname: Liu
authors.affiliation: Jiangsu Provincial Key Laboratory of Radiation Medicine and Protection, School of Radiation Medicine and Protection, Medical College of Soochow University, Suzhou, Jiangsu 215123, P.R. China.
authors.forename: Feng-Mei
authors.initials: FM
authors.lastname: Cui
authors.affiliation: Jiangsu Provincial Key Laboratory of Radiation Medicine and Protection, School of Radiation Medicine and Protection, Medical College of Soochow University, Suzhou, Jiangsu 215123, P.R. China.
authors.forename: Jia-Ying
authors.initials: JY
authors.lastname: Xu
authors.affiliation: School of Public Health, Medical College of Soochow University, Suzhou, Jiangsu 215123, P.R. China.
authors.forename: Jian
authors.initials: J
authors.lastname: Tong
authors.affiliation: Department of Radiotherapy, The First Affiliated Hospital of Soochow University, Suzhou, Jiangsu 215006, P.R. China.
authors.forename: Xiao-Fei
authors.initials: XF
authors.lastname: Qi
authors.affiliation: Department of Urology, The First Affiliated Hospital of Soochow University, Suzhou, Jiangsu 215006, P.R. China.
authors.forename: Li-Li
authors.initials: LL
authors.lastname: Wang
authors.affiliation: Jiangsu Provincial Key Laboratory of Radiation Medicine and Protection, School of Radiation Medicine and Protection, Medical College of Soochow University, Suzhou, Jiangsu 215123, P.R. China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhu
keywords: DNA damage response; cell cycle; long non-coding RNA; p53; radiation-induced lung injury; radiosensitivity