Induced expression of the IER5 gene by γ-ray irradiation and its involvement in cell cycle checkpoint control and survival
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The immediate-early response gene 5 (IER5) was previously shown, using microarray analysis, to be upregulated by ionizing radiation. Here we further characterized the dose- and time-dependency of radiation-induced expression of IER5 at doses from 0.5 to 15 Gy by quantitative real-time PCR analyses in HeLa cells and human lymphoblastoid AHH-1 cells. A radiation-induced increase in the IER5 mRNA level was evident 2 h after irradiation with 2 Gy in both cell lines. In AHH-1 cells the expression reached a peak at 4 h and then quickly returned to the control level, while in HeLa cells the expression only remained increased for a short period of time at around 2 h after irradiation before returning to the control. After high-dose irradiation (10 Gy), the induction of the IER5 expression was lower and delayed in AHH-1 cells as compared with 2-Gy irradiated cells. In HeLa cells, at this dose, two peaks of increased expression were observed 2 h and 12–24 h post-irradiation, respectively. RNA interference technology was employed to silence the IER5 gene in HeLa cells. siRNA-mediated suppression of IER5 resulted in an increased proliferation of HeLa cells. Cell growth and survival analyses demonstrated that suppression of IER5 significantly increased the radioresistance of HeLa cells to radiation doses of up to 6 Gy, but barely affected the sensitivity of cells at 8 Gy. Moreover, suppression of IER5 potentiated radiation-induced arrest at the G2-M transition and led to an increase in the fraction of S phase cells. Taken together, we propose that the early radiation-induced expression of IER5 affects the radiosensitivity via disturbing radiation-induced cell cycle checkpoints.
KeywordsHeLa Cell Cell Cycle Checkpoint Clonogenic Survival Assay Flow Cytometry Immunofluorescence Primary Human Fibroblast Cell
This work was supported by grants from the National Basic Research Program of MOST, China (973 Program, Grant No: 2007CB914603), the National Natural Science Foundation of China (Grant No: 30371232, No.:30770533), and the Beijing Municipal Education Commission (Science and Technology for Development Program: Km200710025007).
- Boerma M, van der Wees CGC, Vrieling H, Svensson JP, Wondergem J, van der Laarse A, Mullenders LHF, van Zeeland AA (2005) Microarray analysis of gene expression profiles of cardiac myocytes and fibroblasts after mechanical stress, ionising or ultraviolet radiation. BMC Genomics 6:6–15CrossRefGoogle Scholar
- Chaudhry MA, Chodosh LA, McKenna WG, Muschel RJ (2003) Gene expression profile of human cells irradiated in G1 and G2 phases of cell cycle. Cancer Lett 195:221–233Google Scholar
- Franken NA, Ten Cate R, van Bree C, Haveman J (2004) Induction of the early response protein EGR-1 in human tumour cells after ionizing radiation is correlated with a reduction of repair of lethal lesions and an increase of repair of sublethal lesions. Int J Oncol 24:1027–1031Google Scholar
- Hsu JY, Sun ZW, Li X, Reuben M, Tatchell K, Bishop DK, Grushcow JM, Brame CJ, Caldwell JA, Hunt DF, Lin F, Smith MM, Allis CD (2000) Mitotic phosphorylation of histone H3 is governed by Ipl1/aurora kinase and Glc7/PP1 phosphatase in budding yeast and nematodes. Cell 102:279–291CrossRefGoogle Scholar
- Kis E, Szatmári T, Keszei M, Farkas R, Esik O, Lumniczky K, Falus A, Sáfrány G (2006) Microarray analysis of radiation response genes in primary human fibroblasts. Int J Radiat Oncol Biol Phys 66:1506–1514Google Scholar
- Long XH, Zhao ZQ, He XP, Wang HP, Xu QZ, An J, Bai B, Sui JL, Zhou PK (2007) Dose-dependent expression changes of early response genes to ionizing radiation in human lymphoblastoid cells. Int J Mol Med 19:607–615Google Scholar
- Wang HP, Long XH, Sun ZZ, Rigaud O, Xu QZ, Huang YC, Sui JL, Bai B, Zhou PK (2006) Identification of differentially transcribed genes in human lymphoblastoid cells irradiated with 0.5 Gy of γ-ray and the involvement of low dose radiation inducible CHD6 gene in cell proliferation and radiosensitivity. Int J Radiat Biol 82:181–190CrossRefGoogle Scholar