Theranostics 2018; 8(5):1286-1300. doi:10.7150/thno.22552

Research Paper

S6K1 phosphorylation-dependent degradation of Mxi1 by β-Trcp ubiquitin ligase promotes Myc activation and radioresistance in lung cancer

Yumei Huang1*, Kaishun Hu2*, Sheng Zhang1*, Xiaorong Dong1, Zhongyuan Yin1, Rui Meng1, Yingchao Zhao1, Xiaofang Dai1, Tao Zhang1, Kunyu Yang1, Li Liu1, Kai Huang3, Shaojun Shi4, Yu Zhang4, Junjie Chen5, Gang Wu1✉, Shuangbing Xu1✉

1. Cancer Center, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China;
2. Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Medical Research Center, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou 510120, China;
3. Clinic Center of Human Gene Research, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China;
4. Department of Pharmacy, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China;
5. Department of Experimental Radiation Oncology, The University of Texas M.D. Anderson Cancer Center, Houston, Texas 77030, USA.
* These authors contributed equally to this work.

Abstract

Rationale: Mxi1 is regarded as a potential tumor suppressor protein that antagonizes the transcriptional activity of proto-oncogene Myc. However, the clinical significances and underlying mechanisms by which Mxi1 is regulated in lung cancer remain poorly understood.

Methods: Mass spectrometry analysis and immunoprecipitation assay were utilized to detect the protein-protein interaction. The phosphorylation of Mxi1 was evaluated by in vitro kinase assays. Poly-ubiquitination of Mxi1 was examined by in vivo ubiquitination assay. Lung cancer cells stably expressing wild-type Mxi1 or Mxi1-S160A were used for functional analyses. The expression levels of Mxi1 and S6K1 were determined by immunohistochemistry in lung cancer tissues and adjacent normal lung tissues.

Results: We found that Mxi1 is downregulated and correlated with poor prognosis in lung cancer. Using tandem affinity purification technology, we provided evidence that β-Trcp E3 ubiquitin ligase interacts with and promotes the ubiquitination and degradation of Mxi1. Furthermore, we demonstrated that Mxi1 is phosphorylated at S160 site by the protein kinase S6K1 and subsequently degraded via the ubiquitin ligase β-Trcp. Moreover, a phosphorylation mutant form of Mxi1 (Mxi1-S160A), which cannot be degraded by S6K1 and β-Trcp, is much more stable and efficient in suppressing the transcriptional activity of Myc and radioresistance in lung cancer cells. More importantly, a strong inverse correlation between S6K1 and Mxi1 expression was observed in human lung cancer tissues.

Conclusion: Our findings not only establish a crosstalk between the mTOR/S6K1 signaling pathway and Myc activation, but also suggest that targeting S6K1/Mxi1 pathway is a promising therapeutic strategy for the treatment of lung cancer.

Keywords: Mxi1, S6K1, β-Trcp, phosphorylation, lung cancer

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How to cite this article:
Huang Y, Hu K, Zhang S, Dong X, Yin Z, Meng R, Zhao Y, Dai X, Zhang T, Yang K, Liu L, Huang K, Shi S, Zhang Y, Chen J, Wu G, Xu S. S6K1 phosphorylation-dependent degradation of Mxi1 by β-Trcp ubiquitin ligase promotes Myc activation and radioresistance in lung cancer. Theranostics 2018; 8(5):1286-1300. doi:10.7150/thno.22552. Available from http://www.thno.org/v08p1286.htm