Int J Med Sci 2017; 14(5):452-461. doi:10.7150/ijms.18329

Research Paper

Stabilization of 4E-BP1 by PI3K kinase and its involvement in CHK2 phosphorylation in the cellular response to radiation

Zi-Jian Yu1, Hui-Hui Luo2, 3, Zeng-Fu Shang4, Hua Guan3, Bei-Bei Xiao4, Xiao-Dan Liu3, Yu Wang3, Bo Huang2✉, Ping-Kun Zhou2, 3✉

1. Department of Hepatobiliary Surgery, the First Affiliated Hospital, University of South China, Hengyang, Hunan Province 421001, P.R. China;
2. Institute for Environmental Medicine and Radiation Health, the College of Public Health, University of South China, Hengyang, Hunan Province 421001, P.R. China;
3. Department of Radiation Toxicology and Oncology, Beijing Key Laboratory for Radiobiology (BKLRB), Beijing Institute of Radiation Medicine, 100850 Beijing, P.R. China;
4. School of Radiation Medicine and Protection, Medical College of Soochow University, Collaborative Innovation Center of Radiation Medicine of Jiangsu Higher Education Institutions, Suzhou, Jiangsu Province 215123, P.R. China.

Abstract

Objectives: 4E-BP1 is a family member of eIF4E binding proteins (4E-BPs) which act as the suppressors of cap-dependent translation of RNA via competitively associating with cap-bound eIF4E. RNA translation regulation is an important manner to control the cellular responses to a series of stress conditions such as ionizing radiation (IR)-induced DNA damage response and cell cycle controlling. This study aimed to determine the mechanism of 4E-BP1 stabilization and its potential downstream target(s) in the response to IR.

Methods: PI3Ks kinase inhibitors were used to determine the signaling control of 4E-BP1 phosphorylation and protein stability. shRNA strategy was employed to silence the expression of 4E-BP1 in HeLa and HepG2 cells, and determine its effect on the irradiation-induced CHK2 phosphorylation. The protein degradation/stability was investigated by western blotting on the condition of blocking novel protein synthesis by cycloheximide (CHX).

Results: The phosphorylation of 4E-BP1 at Thr37/46 was significantly increased in both HepG2 and HeLa cells by ionizing radiation. Depression of 4E-BP1 by shRNA strategy resulted in an incomplete G2 arrest at the early stage of 2 hours post-irradiation, as well as a higher accumulation of mitotic cells at 10 and 12 hours post-irradiation as compared to the control cells. Consistently, the CHK2 phosphorylation at Thr68 induced by IR was also attenuated by silencing 4E-BP1 expression. Both PI3K and DNA-PKcs kinase inhibitors significantly decreased the protein level of 4E-BP1, which was associated with the accelerated degradation mediated by ubiquitination-proteasome pathway.

Conclusion: PI3K kinase activity is necessary for maintaining 4E-BP1 stability. Our results also suggest 4E-BP1 a novel biological role of regulating cell cycle G2 checkpoint in responding to IR stress in association with controlling CHK2 phosphorylation.

Keywords: 4E-BP1, PI3K kinase

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How to cite this article:
Yu ZJ, Luo HH, Shang ZF, Guan H, Xiao BB, Liu XD, Wang Y, Huang B, Zhou PK. Stabilization of 4E-BP1 by PI3K kinase and its involvement in CHK2 phosphorylation in the cellular response to radiation. Int J Med Sci 2017; 14(5):452-461. doi:10.7150/ijms.18329. Available from http://www.medsci.org/v14p0452.htm