Cancer Letters

Cancer Letters

Volume 436, 1 November 2018, Pages 139-148
Cancer Letters

Original Articles
USP10 suppresses tumor progression by inhibiting mTOR activation in hepatocellular carcinoma

https://doi.org/10.1016/j.canlet.2018.07.032Get rights and content

Highlights

  • USP10 is significantly downregulated and associated with poor prognosis in HCC.

  • USP10 stabilizes PTEN and AMPKα by inhibiting PTEN and AMPKα polyubiquitylation.

  • USP10 inhibits hepatocellular carcinoma growth in vivo by inhibiting mTOR signaling pathway.

Abstract

Dysregulation of deubiquitination pathway is associated with poor prognosis in cancers such as hepatocellular carcinoma (HCC). The mammalian target of rapamycin, mTOR, has become an attractive cancer therapeutic target in HCC. However, whether and how aberrant expression of deubiquitination pathway regulates mTOR pathway has remained elusive. Here we report that ubiquitin-specific protease 10 (USP10) functions as a tumor suppressor which inhibits mTOR pathway by stabilizing PTEN and AMPKα in HCC cells. Mechanistically, USP10 interacts and stabilizes PTEN and AMPKα by inhibiting their polyubiquitylation. This stabilization in turn inhibits AKT phosphorylation and mTOR Complex1 (mTORC1) activation. In human liver cancer, USP10 expression is downregulated in HCC tumor tissues across three independent HCC cohorts, and lower-expression of USP10 will generate poor prognosis outcome. Collectively, our results uncover an undescribed mechanism where USP10, as a tumor suppressor, negatively regulates mTORC1 activation and AKT phosphorylation by stabilizing AMPKα and PTEN in HCC cells. This study sheds light on the theoretical basis of mTOR signaling pathway-oriented targeting treatment in clinic.

Introduction

The mammalian target of rapamycine, mTOR, belongs to phosphoinositide 3-kinase (PI3K)-related kinase family. As a central regulator, it controls cell growth, proliferation, differentiation and survival [1]. Hyperactivation of mTOR signaling is frequently found in different types of cancer such as HCC [2]. Targeting on this pathway provides an attractive strategy for improving the clinical therapeutic effects of HCC patients. However, due to the complexity and heterogeneity of HCC cells, single agent targeting therapy on mTOR does not reach an expected clinical efficacy [3]. Therefore, discovering the regulatory proteins of mTOR signaling pathway is important for improving cancer care in HCC.

PTEN/PI3K/AKT and LKB1/AMPKα signaling pathways are well-established upstream regulators of mTOR activation. PTEN, as an antagonizer of PI3K/AKT signaling, plays critical roles in many cellular processes [[4], [5], [6]]. AKT directly phosphorylates tuberous sclerosis 2 (TSC2), which impairs the ability of TSC2 to inhibit mTORC1 activation [7,8]. On the other hand, AMPKα is activated by LKB1 and energy homeostasis mediated phosphorylation and then phosphorylates TSC2 to inhibit mTORC1 function in cancer cells [9,10]. Thus, establishment of a fine tuning factor which inhibits mTOR activation through PTEN/PI3K/AKT and LKB1/AMPK signaling pathways may be an alternative way to develop anticancer mTOR inhibitors and improve clinical outcomes in cancer therapies.

Ubiquitination and deubiquitination, two reversal processes, are acknowledged as important post-translational modifications which regulate protein degradation, localization and activation in cells by conjugating or removing the ubiquitin from substrate proteins [11]. Dysregulation of ubiquitination and deubiquitination contributed to oncogenic progression in several types of cancer [[12], [13], [14]]. As previous studies have reported, USP10 regulates crucial signaling factors involved in cell proliferation, apoptosis, autophagy and cancer metabolism, such as p53 [15], PTEN [16], TANK [17], SIRT6 [18], MSH2 [19], and autophagy related protein Beclin1 [20].

In this study, we investigated on the crucial signaling regulatory functions of USP10 and identified USP10 as a tumor suppressor in HCC cells. USP10 expression is significantly downregulated in HCC tumors and lower-expression of USP10 will lead to worse prognosis in HCC cohorts. Mechanistically, USP10 interacts with and stabilizes PTEN and AMPKα by inhibiting their polyubiquitylation, and subsequently suppresses the activity of mTOR signaling pathway. Collectively, these results indicate that USP10 may be a new therapeutic target in mTOR signaling activated HCC cells.

Section snippets

Cell culture

The immortalized human liver cell line THLE2 was purchased from the American Type Culture Collection (ATCC, Manassas, VA, USA). The human HCC cell lines SUN-449, BEL-7402, SMMC-7721, HepG2, HUH7, MHCC-97H, MHCC-97 L, HCCLM3 and HB611 were purchased from the cell bank of the Committee on Type Culture Collection of the Chinese Academy of Sciences (CTCC, Shanghai, China). THLE2 cells were cultured in Bronchial Epithelial Cell Growth Medium (BEGM, Lonza/Clonetics Corporation, Walkersville)

USP10 expression is downregulated and associated with poor prognosis in HCC

To determine whether USP10 is dysregulated in HCC tissues, we performed immunoblotting and immunohistochemistry analyses in human HCC tissues. We observed that USP10 was significantly downregulated in HCC tissues compared with adjacent non-cancer tissues (Fig. 1A, B and S1A, B). Additionally, this observation was further confirmed by immunohistochemical staining of USP10 in two representative HCC and adjacent non-tumor specimens (Fig. 1C) and 74 pairs of paraffin-embedded tissues (Fig. 1D).

Discussion

To date, the role of the deubiquitinating enzyme USP10 in different types of cancer is still controversy. Previously, it has been reported that USP10 plays as a tumor initiator in prostate cancer [28], FLT3-ITD-positive AML [29] and glioblastoma multiforme [30], whereas it also exhibits as a tumor suppressor in lung cancer [16], gastric cancer [31], colon cancer [18] and renal cell carcinoma [15]. However, the significant mechanistic role and expression of USP10 in HCC remain largely unclear.

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.

Author contributions

N.Z., C.L. and H-l.P. conceived the project, and H-l.P. and G.T. supervised the project. N.Z., C.L., A.W. and D.C. designed and performed most of experiments and the data analysis. X.L.T.X., D.S.T., W.W., T.L., X.L., J.L., H.Q., H.L, J.D., C.M., G.X., Q.Y. and J.L. provided significant intellectual input. N.Z., G.T., D.C. and H-l.P. wrote the manuscript with input from all other authors.

Acknowledgements

We thank members of the Dr. Piao laboratory for helpful discussion. This study is supported by the National Key Research and Development Program of China (2016YFC0903302), National Natural Science Foundation of China (No. 81502024, No. 81672440, No. 31701156), Innovation program of science and research from the DICP, CAS (DICP TMSR201601) and the 100 Talents Program of Chinese Academy of Sciences.

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