Stabilization of histone demethylase PHF8 by USP7 promotes breast carcinogenesis
Qian Wang, … , Zhi Yao, Lei Shi
Qian Wang, … , Zhi Yao, Lei Shi
Published May 16, 2016
Citation Information: J Clin Invest. 2016;126(6):2205-2220. https://doi.org/10.1172/JCI85747.
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Research Article Oncology

Stabilization of histone demethylase PHF8 by USP7 promotes breast carcinogenesis

Abstract

The histone demethylase PHF8 has been implicated in multiple pathological disorders, including X-linked mental retardation and tumorigenesis. However, it is not clear how the abundance and function of PHF8 are regulated. Here, we report that PHF8 physically associates with the deubiquitinase USP7. Specifically, we demonstrated that USP7 promotes deubiquitination and stabilization of PHF8, leading to the upregulation of a group of genes, including cyclin A2, that are critical for cell growth and proliferation. The USP7-encoding gene was also transcriptionally regulated by PHF8, via positive feedback. USP7 was overexpressed in breast carcinomas, and the level of expression positively correlated with expression of PHF8 and cyclin A2 and with the histological grade of breast cancer. We showed that USP7 promotes breast carcinogenesis by stabilizing PHF8 and upregulating cyclin A2 and that the interaction between USP7 and PHF8 is augmented during DNA damage. Moreover, USP7-promoted PHF8 stabilization conferred cellular resistance to genotoxic insults and was required for the recruitment of BLM and KU70, which are both essential for DNA double-strand break repair. Our study mechanistically links USP7 to epigenetic regulation and DNA repair. Moreover, these data support the pursuit of USP7 and PHF8 as potential targets for breast cancer intervention, especially in combination with chemo- or radiotherapies.

Authors

Qian Wang, Shuai Ma, Nan Song, Xin Li, Ling Liu, Shangda Yang, Xiang Ding, Lin Shan, Xing Zhou, Dongxue Su, Yue Wang, Qi Zhang, Xinhua Liu, Na Yu, Kai Zhang, Yongfeng Shang, Zhi Yao, Lei Shi

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Figure 8

PHF8 promotes BLM and KU70 accumulation at DSBs.

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PHF8 promotes BLM and KU70 accumulation at DSBs. (A) MCF-7 cells stably ...
(A) MCF-7 cells stably expressing control vector, PHF8/WT, or PHF8/H247A were transfected with the indicated siRNAs and subjected to UV-laser microdissection. The cells were fixed and immunostained with anti-γH2AX and anti-BLM followed by microscopy analysis. Scale bar: 10 μm. Percentage of nuclei with double stripes was quantified, and each bar represents the mean ± SD for biological triplicate experiments with more than 100 nuclei. Relative intensities of BLM stripes to the uncut regions determined by ImageJ software are presented with box plots. The circle indicates the outlier value determined by SPSS software. (B) Whole-cell lysates from MCF-7 cells (left panel) or MCF-7 cells stably expressing PHF8/WT or PHF8/H247A (right panel) were immunoprecipitated and then immunoblotted with antibodies against the indicated proteins. (C) MCF-7 cells stably expressing control vector, PHF8/WT, or PHF8/H247A were cotransfected with the indicated siRNAs and GFP-KU70 followed by UV-laser microirradiation. Scale bar: 10 μm. Relative fluorescence intensities in microirradiated areas to the undamaged regions were determined by ImageJ software. More than 20 nuclei were scored in biological triplicate experiments. Each bar represents the mean ± SD. *P < 0.05 and **P < 0.01, 1-way ANOVA in A and 2-way ANOVA in C. pLenti, lentiviral plasmid.