Ubiquitin-specific peptidase 14 maintains estrogen receptor α stability via its deubiquitination activity in endometrial cancer

Abstract

USP14 deubiquitinates ERα to maintain its stability in ECEndometrial cancer (EC) is one of the common gynecological malignancies of which the incidence has been rising for decades. It is considered that continuously unopposed estrogen exposure is the main risk factor for EC initiation. Thus, exploring the modulation of estrogen/estrogen receptor α (ERα) signaling pathway in EC would be helpful to well understand the mechanism of EC development and find the potential target for EC therapy. Ubiquitin-specific peptidase 14 (USP14), a member of the proteasome-associated deubiquitinating enzyme family, plays a crucial role in a series of tumors. However, the function of USP14 in EC is still elusive. Here, our results have demonstrated that USP14 is highly expressed in EC tissues compared with that in normal endometrial tissues, and higher expression of USP14 is positively correlated with poor prognosis. Moreover, USP14 maintains ERα stability through its deubiquitination activity. Our results further demonstrate that USP14 depletion decreases the expression of ERα-regulated genes in EC-derived cell lines. Moreover, knockdown of USP14 or USP14-specific inhibitor treatment significantly suppresses cell growth and migration in EC cell lines or in mice. We further provide the evidence to show that the effect of USP14 on EC cell growth, if not all, at least is partially related to ERα pathway. Our study provides new sights for USP14 to be a potential therapeutic target for the treatment of EC, especially for EC patients with fertility preservation needs. USP14 deubiquitinates ERα to maintain its stability in ECEndometrial cancer (EC) is one of the common gynecological malignancies of which the incidence has been rising for decades. It is considered that continuously unopposed estrogen exposure is the main risk factor for EC initiation. Thus, exploring the modulation of estrogen/estrogen receptor α (ERα) signaling pathway in EC would be helpful to well understand the mechanism of EC development and find the potential target for EC therapy. Ubiquitin-specific peptidase 14 (USP14), a member of the proteasome-associated deubiquitinating enzyme family, plays a crucial role in a series of tumors. However, the function of USP14 in EC is still elusive. Here, our results have demonstrated that USP14 is highly expressed in EC tissues compared with that in normal endometrial tissues, and higher expression of USP14 is positively correlated with poor prognosis. Moreover, USP14 maintains ERα stability through its deubiquitination activity. Our results further demonstrate that USP14 depletion decreases the expression of ERα-regulated genes in EC-derived cell lines. Moreover, knockdown of USP14 or USP14-specific inhibitor treatment significantly suppresses cell growth and migration in EC cell lines or in mice. We further provide the evidence to show that the effect of USP14 on EC cell growth, if not all, at least is partially related to ERα pathway. Our study provides new sights for USP14 to be a potential therapeutic target for the treatment of EC, especially for EC patients with fertility preservation needs. Endometrial cancer (EC) is one of the gynecological malignancies, with an increasing incidence in the world (1Ferlay J. Soerjomataram I. Dikshit R. Eser S. Mathers C. Rebelo M. et al.Cancer incidence and mortality worldwide: sources, methods and major patterns in GLOBOCAN 2012.Int. J. Cancer. 2015; 136: E359-E386Crossref PubMed Scopus (21762) Google Scholar, 2Bray F. Ferlay J. Soerjomataram I. Siegel R.L. Torre L.A. Jemal A. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries.CA Cancer J. Clin. 2018; 68: 394-424Crossref PubMed Scopus (53408) Google Scholar). It has been reported that EC patients who are at advanced stage or suffer from recurrence have poor prognosis (3Arend R.C. Jones B.A. Martinez A. Goodfellow P. Endometrial cancer: molecular markers and management of advanced stage disease.Gynecol. Oncol. 2018; 150: 569-580Abstract Full Text Full Text PDF PubMed Scopus (109) Google Scholar). According to the updated statistics, there are about 66,570 new cases and 12,940 deaths in the United States in 2021 (4Siegel R.L. Miller K.D. Fuchs H.E. Jemal A. Cancer statistics, 2021.CA Cancer J. Clin. 2021; 71: 7-33Crossref PubMed Scopus (10389) Google Scholar), indicating an increase in the morbidity and mortality of EC. Importantly, because of the younger age of EC onset, the treatment strategies with fertility preservation are critical and challenging. Therefore, it is necessary to understand the molecular mechanism of EC development and explore more effective and therapeutic target for EC treatment. In consideration of hormonal dependence in EC, it is divided into two types, including type I hormone dependent and type II hormone independent (5Bokhman J.V. Two pathogenetic types of endometrial carcinoma.Gynecol. Oncol. 1983; 15: 10-17Abstract Full Text PDF PubMed Scopus (1864) Google Scholar). Type I accounts for 80% of the EC cases, with the feature of positive estrogen receptor α (ERα) expression (6Carlson M.J. Thiel K.W. Leslie K.K. Past, present, and future of hormonal therapy in recurrent endometrial cancer.Int. J. Womens Health. 2014; 6: 429-435PubMed Google Scholar). It is generally known that long-term unopposed estrogen (E2) exposure is the main risk factor for type I EC, indicating that E2–ERα signaling pathway plays a significant role in EC development and progression. Previous studies have shown that ERα, as a transcription factor, is involved in the pathogenic process in breast cancer (BCa) (7Tecalco-Cruz A.C. Ramirez-Jarquin J.O. Cruz-Ramos E. Estrogen receptor alpha and its ubiquitination in breast cancer cells.Curr. Drug Targets. 2019; 20: 690-704Crossref PubMed Scopus (19) Google Scholar). A number of coregulators involved in the modulation of ERα activity promote tumorigenesis and contribute to endocrine therapy resistance in BCa (8Xia X. Liao Y. Huang C. Liu Y. He J. Shao Z. et al.Deubiquitination and stabilization of estrogen receptor alpha by ubiquitin-specific protease 7 promotes breast tumorigenesis.Cancer Lett. 2019; 465: 118-128Crossref PubMed Scopus (54) Google Scholar, 9Tang J. Luo Y. Tian Z. Liao X. Cui Q. Yang Q. et al.TRIM11 promotes breast cancer cell proliferation by stabilizing estrogen receptor alpha.Neoplasia. 2020; 22: 343-351Crossref PubMed Scopus (28) Google Scholar, 10Chen X.S. Wang K.S. Guo W. Li L.Y. Yu P. Sun X.Y. et al.UCH-L1-mediated down-regulation of estrogen receptor alpha contributes to insensitivity to endocrine therapy for breast cancer.Theranostics. 2020; 10: 1833-1848Crossref PubMed Scopus (18) Google Scholar). However, the regulation of ERα signaling pathway and its underlying biological function in EC progression are still largely unknown. Ubiquitin-specific protease 14 (USP14), as one of the proteasome-associated deubiquitinase, exerts important roles in several carcinomas (11Yu F. Liu J.B. Wu Z.J. Xie W.T. Zhong X.J. Hou L.K. et al.Tumor suppressive microRNA-124a inhibits stemness and enhances gefitinib sensitivity of non-small cell lung cancer cells by targeting ubiquitin-specific protease 14.Cancer Lett. 2018; 427: 74-84Crossref PubMed Scopus (40) Google Scholar, 12Sharma A. Almasan A. USP14 regulates DNA damage response and is a target for radiosensitization in non-small cell lung cancer.Int. J. Mol. Sci. 2020; 21: 6383Crossref PubMed Scopus (13) Google Scholar). It has been reported that USP14 appears to have quite different functions in modulating intracellular proteolytic degradation involving in both maintenance of protein stability and protein degradation (13Liu B. Jiang S. Li M. Xiong X. Zhu M. Li D. et al.Proteome-wide analysis of USP14 substrates revealed its role in hepatosteatosis via stabilization of FASN.Nat. Commun. 2018; 9: 4770Crossref PubMed Scopus (61) Google Scholar, 14Kim H.T. Goldberg A.L. UBL domain of Usp14 and other proteins stimulates proteasome activities and protein degradation in cells.Proc. Natl. Acad. Sci. U. S. A. 2018; 115: E11642-E11650Crossref PubMed Scopus (38) Google Scholar). Ubp6, a yeast homolog of USP14, participates in delaying protein degradation to lead to protein accumulation (15Hanna J. Hathaway N.A. Tone Y. Crosas B. Elsasser S. Kirkpatrick D.S. et al.Deubiquitinating enzyme Ubp6 functions noncatalytically to delay proteasomal degradation.Cell. 2006; 127: 99-111Abstract Full Text Full Text PDF PubMed Scopus (279) Google Scholar). Studies have been shown that USP14 stabilizes androgen receptor (AR) protein through its deubiquitination activity. USP14 can also promote cell growth and inhibit cell apoptosis in AR-positive ERα-negative BCa (16Liao Y. Xia X. Liu N. Cai J. Guo Z. Li Y. et al.Growth arrest and apoptosis induction in androgen receptor-positive human breast cancer cells by inhibition of USP14-mediated androgen receptor deubiquitination.Oncogene. 2018; 37: 1896-1910Crossref PubMed Scopus (76) Google Scholar). USP14 interacts with murine double minute 2 and stabilizes murine double minute 2, leading to tumor progression in cervical cancer (17Xu L. Wang J. Yuan X. Yang S. Xu X. Li K. et al.IU1 suppresses proliferation of cervical cancer cells through MDM2 degradation.Int. J. Biol. Sci. 2020; 16: 2951-2963Crossref PubMed Scopus (13) Google Scholar). USP14 regulates cisplatin resistance in ovarian cancer through stabilizing BCL6 protein (18Shen J. Hong L. Chen L. Ubiquitin-specific protease 14 regulates ovarian cancer cisplatin-resistance by stabilizing BCL6 oncoprotein.Biochem. Biophysical Res. Commun. 2020; 524: 683-688Crossref PubMed Scopus (9) Google Scholar). However, the functional analysis of USP14 in EC is still largely elusive. In this study, we have demonstrated that USP14 is highly expressed in EC clinical samples compared with that in noncarcinoma endometrial tissues. The higher expression level of USP14 is positively correlated with the poor prognosis of EC. Our results also provide the evidence that USP14 is involved in the maintenance of ERα stability through its deubiquitinating activity. USP14 deubiquitinates ERα at lysine 48 (K48)-linked ubiquitination. In addition, depletion of USP14 downregulates ERα-induced gene transcription. Knockdown of USP14 decreases the recruitment of ERα to estrogen-response element (ERE) of c-Myc, which is one of the putative ERα target genes. Furthermore, USP14 depletion or USP14-specific inhibitor attenuates cell growth and migration in EC-derived cell lines or in mice. Taken together, our study may provide a potential therapeutic target for the treatment of EC, especially for EC patients with fertility preservation needs. Previous studies have shown that USP14 is highly expressed in a variety of malignancies and plays a critical role in tumor progression (19Wang D. Ma H. Zhao Y. Zhao J. Ubiquitin-specific protease 14 is a new therapeutic target for the treatment of diseases.J. Cell Physiol. 2021; 236: 3396-3405Crossref PubMed Scopus (14) Google Scholar). However, the expression level and functions of USP14 in EC remain poorly defined. To this end, we then analyzed the expression of USP14 in EC tissues and normal endometrial tissues using UALCAN (http://ualcan.path.uab.edu/index.html). The results demonstrated that USP14 has a higher expression in EC tissues (Fig. D.S. B. I. B. et a for tumor gene expression and PubMed Scopus Google Scholar). We then analyzed the expression level of USP14 at different clinical and The results that there the and (Fig. A and To further there a USP14 and the clinical of the we an analysis through on the Cancer A. B. analysis of cancer 2021; PubMed Scopus Google the results that the expression of USP14 with the and (Fig. and We then the expression of USP14 in the normal tissues and EC tissues from the of to further the expression of USP14 in noncarcinoma tissues and EC tissues. The results from or that the expression level of USP14 significantly higher in the EC tissues compared with that in the tissues (Fig. that ERα plays a significant role in EC we then the expression of USP14 and ERα in EC Our results that the expression of USP14 positively correlated with that of ERα in EC samples (Fig. and known the the expression of USP14 and ERα, we then to the underlying mechanism We the expression of ERα expression of USP14, which USP14 ERα at protein level in a and in the EC cell lines with USP14 depletion or with a inhibitor of USP14 M.J. S. Elsasser S. Chen et of proteasome activity by a inhibitor of PubMed Scopus Google Scholar), a significant in ERα expression at the protein level not at the level 2, and and results that USP14 ERα by its degradation not its transcription. To further this we the cells with protein to the of USP14 on ERα in human and human endometrial The results demonstrated that USP14 ERα inhibition of USP14 ERα degradation (Fig. 2, In addition, in the of the proteasome inhibitor the increase of ERα by USP14 in both and cell lines 2, and Taken together, that USP14 be involved in the maintenance of ERα stability via the proteasome pathway in EC cell lines. We further to USP14 interacts with The expression of and then to in cell lines. The results demonstrated that USP14 with ERα with or treatment (Fig. To further we in EC cell lines and cell the results an USP14 and ERα in EC-derived cell lines (Fig. and of the deubiquitination activity of USP14, we USP14 can in of ERα ubiquitination. ubiquitination then as The results demonstrated that the expression of USP14 the level of ERα ubiquitination in EC cell lines (Fig. Moreover, USP14 depletion or USP14 inhibitor treatment significantly ERα ubiquitination (Fig. and Thus, results that USP14 in deubiquitination of ERα to maintain stabilization of ERα It has been well that different types of protein of which and lysine a for proteasomal degradation Y. A. The proteasomal not a PubMed Scopus Google Scholar, H.T. F. D. et of and Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). We to which type of on ERα by Our results that expression of USP14 ubiquitination of ERα not ubiquitination in cells (Fig. Furthermore, different of the expression of USP14 a significant of ubiquitination of ERα in a in cells (Fig. all, our results that USP14 involved in ERα deubiquitination with to maintain the stability of that USP14 may increase the accumulation of ERα, we then to study the genes of ERα by USP14 in EC-derived to the expression level of ERα target genes. We the level of and in EC-derived cell lines with knockdown of Our results that depletion of USP14 different of and the expression of ERα-regulated genes in the of in cells (Fig. To further this the cells with USP14 expression with of as The that the of USP14 expression on ERα-regulated genes significantly by treatment (Fig. We further the protein expression of ERα genes by and our demonstrated that depletion of USP14 the protein expression of ERα-regulated genes. expression of USP14 protein of ERα-regulated genes as in EC-derived cells (Fig. and We then to USP14 on the recruitment of ERα on the of ERα target we to the putative which the of on the of ERα-regulated as (Fig. then to the recruitment of ERα at the in the cells USP14 The depletion of USP14 shown to the recruitment of ERα on the of gene (Fig. Taken together, USP14 involved in of the transcription of ERα-regulated and USP14 also the recruitment of ERα on the of ERα-regulated To further the function of USP14 in EC-derived we the knockdown of USP14 in and cells The of USP14 knockdown by in EC-derived cell lines with (Fig. in which that the depletion of USP14 inhibit the of Moreover, cells and cells with different of USP14-specific and The results that the in a in EC-derived cell lines and A and We then further USP14 the cell and the analysis in cells with The results that arrest (Fig. and To study the effect of USP14 on the cell proliferation in EC-derived and in the or the of in and Our demonstrated that USP14 cell proliferation in EC-derived cell lines and and In addition, to ERα is for the biological function of USP14 on cell growth in EC-derived of ERα into the cells Our results demonstrated that expression of ERα partially the growth inhibition by (Fig. Taken together, USP14 is involved in of cell growth in EC-derived cells at least partially via To the effect of USP14 on cell the in cells or treatment. The results that inhibition of USP14 significantly the cell migration in cells (Fig. A and Furthermore, with different of the results from the that the migration of cells by treatment in a (Fig. and the further with cells the treatment of The results demonstrated that USP14 depletion cell and expression of USP14 cell migration in the of (Fig. To further the biological function of USP14 in cell growth in we tumor in to the effect of USP14 depletion on cell growth in EC-derived The cells with or into the and of the mice. The results demonstrated that the of and to show the activity. The with or with in (Fig. The tumor and significantly the treatment of or and more the treatment of and (Fig. and then to the expression level of USP14 in tumor from cells (Fig. In addition, to the protein level of USP14, ERα, and in the The results further demonstrated that USP14 depletion significantly ERα and expression (Fig. Taken together, our results that knockdown of USP14 or treatment inhibits EC-derived cell growth in mice. EC is one of the common carcinomas in The treatment of is in patients at an However, the for the patients and the advanced stage patients is EC to one of the the endocrine therapy in EC is not effective compared with that in It has been considered that E2–ERα signaling pathway is for the development of EC. However, the biological function of ERα in EC is still elusive. In this study, our results have demonstrated that USP14 is involved in the maintenance of ERα stabilization to contribute to the progression of EC, a potential therapeutic target for EC especially for fertility preservation of EC patients (Fig. It has been reported that USP14 is highly related to cancer and progression in of one of the proteasome-associated deubiquitinating USP14 has a function in protein degradation and proteasome USP14 is in cancer and AR-positive and the expression of USP14 is positively correlated with that of of USP14 may cell and promote cell apoptosis via stabilizing in cancer or AR-positive BCa (16Liao Y. Xia X. Liu N. Cai J. Guo Z. Li Y. et al.Growth arrest and apoptosis induction in androgen receptor-positive human breast cancer cells by inhibition of USP14-mediated androgen receptor deubiquitination.Oncogene. 2018; 37: 1896-1910Crossref PubMed Scopus (76) Google Scholar, Y. Liu N. X. Cai J. Xia X. Wang X. et ubiquitin-specific protease 14 regulates cancer proliferation by deubiquitinating and stabilizing androgen Scopus Google Scholar). In addition, USP14 enhances signaling pathway via stabilizing to promote tumor development in lung cancer and cancer Li L. W. USP14 promotes tumor progression in 2015; PubMed Scopus (38) Google Scholar, N. Liu C. C. Li Q. of deubiquitinating enzyme USP14 in lung promotes proliferation through the accumulation of J. Mol. Sci. PubMed Scopus Google Scholar). However, the biological function and underlying molecular mechanism of USP14 in EC progression is still Previous study has that USP14 is a of recurrence for EC and inhibition of USP14 with may increase the EC response W. A. S. Zhao X. et is a of recurrence in endometrial cancer and a molecular target for endometrial cancer PubMed Scopus Google Scholar). biological function results are in our Our results that USP14 is in EC clinical and the expression of USP14 is positively correlated with the poor prognosis of EC from our study more on into the USP14 and which is a main risk factor for the of EC. Our study has demonstrated that USP14 is involved in the maintenance of ERα through ERα Moreover, we also provide the evidence that USP14 depletion or USP14 inhibitor treatment cell growth in EC-derived cell lines and that the USP14 and ERα may promote EC progression. estrogen exposure is one of the main risk for type I EC. It has been considered that 80% of type I EC is ERα indicating that modulation of E2–ERα signaling pathway would be for the development of as BCa and EC. It is well known that ERα protein stability is by proteasome studies have that as and may ERα to ERα and then promote BCa progression J. Luo Y. Tian Z. Liao X. Cui Q. Yang Q. et al.TRIM11 promotes breast cancer cell proliferation by stabilizing estrogen receptor alpha.Neoplasia. 2020; 22: 343-351Crossref PubMed Scopus (28) Google Scholar, S. Luo H. Wang C. Sun H. Sun Sun N. et as a of estrogen receptor alpha promotes cell growth in breast Mol. PubMed Scopus Google Scholar). has been to ERα with both and to the stability of ERα, cell growth and endocrine resistance in BCa S. Zhong X. Wang C. Luo H. L. Sun H. et positively via its activity in breast 2020; PubMed Scopus (19) Google Scholar). a deubiquitinase, suppresses ERα transcription through the stability of growth factor which is a of ERα X.S. Wang K.S. Guo W. Li L.Y. Yu P. Sun X.Y. et al.UCH-L1-mediated down-regulation of estrogen receptor alpha contributes to insensitivity to endocrine therapy for breast cancer.Theranostics. 2020; 10: 1833-1848Crossref PubMed Scopus (18) Google Scholar). Thus, well the modulation of ERα in would provide the potential therapeutic for tumor treatment. In this study, our results have shown that USP14 with ERα to maintain the stability of ERα via its deubiquitination activity in EC. USP14 depletion or USP14 inhibitor treatment enhances ERα level with not Our that USP14 involved in the of ERα stability in EC may be a new for the endocrine therapeutic target for EC treatment. Our study has demonstrated that as a of ERα is with to ERα and in the of EC progression K. Wu Y. Wang C. Wang S. Sun H. R. et is involved in of endometrial cancer progression via of Sci. 2020; PubMed Scopus Google Scholar). In addition, it has been reported that in the domain of ERα may of the ERα signaling pathway in EC Goodfellow D. et as a in endometrial Oncol. Full Text Full Text PDF PubMed Scopus Google Scholar). Our have demonstrated that USP14 enhances the recruitment of ERα on the of ERα-regulated increasing the transcription of as and on (Fig. has been as an ERα target gene to promote in EC Y. Zhao R. S. W. C. X. et is by estrogen and promotes via in endometrial Cancer Res. 2020; PubMed Scopus Google Scholar). In with our results have shown that also by USP14 in EC. Taken together, our results that USP14 as the of ERα, the development of EC. In this study has demonstrated that USP14 with ERα to maintain the stability of ERα through its deubiquitination activity with the USP14 to an of ERα recruitment on of ERα target ERα Moreover, depletion of USP14 or USP14-specific inhibitor treatment inhibits cell proliferation and migration in EC, indicating that USP14 may be a therapeutic target in EC treatment. The human EC cell lines and in and cells in the and at with in the USP14 and from from from from The in our and cells with and with DNA to the cells with for 48 The 48 The with for and then with protein for The with as the analysis by the process in our study W. D. Zhao C. M. Wang Y. Ma H. et of signaling in human cancer cells using Res. Commun. PubMed Scopus Google Scholar). with or for for with and DNA using with on the in our in for the genes in in a new from and the from EC cells with to the to our study S. Zhong X. Wang C. Sun H. Wang S. et androgen receptor and promotes cancer Res. PubMed Scopus Google Scholar). cells for with for of by at by with or A to the and then the samples with and the and The DNA samples then by The of cell growth or cells in and with or The cells at and at an of with and cells with The cells in an of at for 14 to the and then into the to the cells for the cells with for or with cells in the with the treatment of different of The of M. The cells of endometrial tissues and EC samples and with the with C. Wang S. L. Wang C. K. Y. et maintains stabilization via its deubiquitination activity in 2021; PubMed Scopus (13) Google Scholar, H.T. Goldberg A.L. The deubiquitinating enzyme Usp14 inhibits proteasomal activities and Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). in the with for at by with of the expression of USP14 and with of the the by an The samples by the of from the by the of cells or into the for The divided into two one with and the other with 2 as X. S. M. Y. Y. Y. et endometrial cancer cells to cisplatin by targeting Lett. 2014; PubMed Scopus Google Scholar). normal and EC tissues from the of the EC samples we for are including stage I cases stage II cases and stage cases the samples to the to the expression level of USP14 and The and with with and considered The are in the main and D.S. B. I. B. et a for tumor gene expression and PubMed Scopus Google Scholar). The that have of with the of this We and for We for the of We also for and for this We and for helpful and K. and Zhao Y. C. S. L. R. R. and B. Z. Y. S. Y. Y. and J. N. Y. C. S. and L. L. and Y. and Zhao study by the of and C. and for of to to and of of to S. by the of for of of and of for and and for of for

Topics & Concepts