Abstract
Interferon-γ (IFN-γ) has been used to control cancers in clinical treatment. However, an increasing number of reports have suggested that in some cases effectiveness declines after a long treatment period, the reason being unclear. We have reported previously that long-term IFN-γ treatment induces malignant transformation of healthy lactating bovine mammary epithelial cells (BMECs) in vitro. In this study, we investigated the mechanisms underlying the malignant proliferation of BMECs under IFN-γ treatment. The primary BMECs used in this study were stimulated by IFN-γ (10 ng/mL) for a long term to promote malignancy. We observed that IFN-γ could promote malignant cell proliferation, increase the expression of cyclin D1/cyclin-dependent kinase 4 (CDK4), decrease the expression of p21, and upregulate the expression of cellular-abelsongene (c-Abl) and histone deacetylase 2 (HDAC2). The HDAC2 inhibitor, valproate (VPA) and the c-Abl inhibitor, imatinib, lowered the expression level of cyclin D1/CDK4, and increased the expression level of p21, leading to an inhibitory effect on IFN-γ-induced malignant cell growth. When c-Abl was downregulated, the HDAC2 level was also decreased by promoted proteasome degradation. These data suggest that IFN-γ promotes the growth of malignant BMECs through the c-Abl/HDAC2 signaling pathway. Our findings suggest that long-term application of IFN-γ may be closely associated with the promotion of cell growth and even the carcinogenesis of breast cancer.
概要
目的
在之前的研究中我们发现γ 干扰素(IFN-γ)通过 营养感受器GCN2 诱导牛乳腺上皮细胞(BMEC) 的恶性转化。在恶性转化的表型中,包括细胞周 期缩短、细胞增殖增加、细胞迁移和侵袭的发生, 而细胞生长的异常调节是细胞恶性转化的第一 步。因此,本研究旨在探讨IFN-γ 诱导细胞恶性 生长的分子机制。
创新点
实验对象γ-BMEC 能够更好的用于乳腺癌发生的 基础研究,是一种新的研究工具。我们选择恶性 细胞生长来详细研究IFN-γ 的作用机制,为 IFN-γ、恶性细胞生长甚至乳腺癌之间的密切关系 提供直接证据。
方法
通过MTT 实验检测IFN-γ 长期刺激下细胞的增殖 能力;蛋白质印迹(Western blot)检测cyclin D1/CDK4、p21、HDAC2、c-Abl 蛋白的表达;免 疫荧光观察c-Abl 入核。
结论
IFN-γ 通过c-Abl/HDAC2 信号通路促进恶性 BMEC 的生长。
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References
Allington TM, Galliher-Beckley AJ, Schiemann WP, 2009. Activated Abl kinase inhibits oncogenic transforming growth factor-β signaling and tumorigenesis in mammary tumors. Faseb J, 23(12):4231–4243. https://doi.org/10.1096/fj.09-13841.
Barneda-Zahonero B, Parra M, 2012. Histone deacetylases and cancer. Mol Oncol, 6(6):579–589. https://doi.org/10.1016/j.molonc.2012.07.003
Bougarn S, Cunha P, Gilbert FB, et al., 2011. Technical note: validation of candidate reference genes for normalization of quantitative PCR in bovine mammary epithelial cells responding to inflammatory stimuli. J Dairy Sci, 94(5): 2425–2430. https://doi.org/10.3168/jds.2010-385.
Bradley WD, Koleske AJ, 2009. Regulation of cell migration and morphogenesis by Abl-family kinases: emerging mechanisms and physiological contexts. J Cell Sci, 122(19):3441–3454. https://doi.org/10.1242/jcs.039859
Brightbill H, Schlissel MS, 2009. The effects of c-Abl mutation on developing B cell differentiation and survival. Int Immunol, 21(5):575–585. https://doi.org/10.1093/intimm/dxp027
Carpi A, Nicolini A, Antonelli A, et al., 2009. Cytokines in the management of high risk or advanced breast cancer: an update and expectation. Curr Cancer Drug Tar, 9(8): 9888–9903. https://doi.org/10.2174/156800909790192392
Creagan ET, Schaid DJ, Ahmann DL, et al., 1990. Disseminated malignant melanoma and recombinant interferon: analysis of seven consecutive phase II investigations. J Invest Dermatol, 95(S6): S188–S192. https://doi.org/10.1111/1523-1747.ep12875512
Gonzalez-Zuñiga M, Contreras PS, Estrada LD, et al., 2014. c-Abl stabilizes HDAC2 levels by tyrosine phosphorylation repressing neuronal gene expression in Alzheimer’s disease. Mol Cell, 56(1):163–173. https://doi.org/10.1016/j.molcel.2014.08.013
Hildmann C, Riester D, Schwienhorst A, 2007. Histone deacetylases—an important class of cellular regulators with a variety of functions. Appl Microbiol Biotechnol, 75(3):487–497. https://doi.org/10.1007/s00253-007-0911-2
Horikoshi T, Fukuzawa KF, Hanada N, et al., 1995. In vitro comparative study of the antitumor effects of human interferon-α, β and γ on the growth and invasive potential of human melanoma cells. J Dermatol, 22(9):631–636. https://doi.org/10.1111/j.1346-8138.1995.tb03889
Kaplan DH, Shankaran V, Dighe AS, et al., 1998. Demonstration of an interferon γ-dependent tumor surveillance system in immunocompetent mice. Proc Natl Acad Sci USA, 95(13):7556–7561. https://doi.org/10.1073/pnas.95.13.7556
Krämer OH, 2009. HDAC2: a critical factor in health and disease. Trends Pharmacol Sci, 30(12):647–655. https://doi.org/10.1016/j.tips.2009.09.007
Krämer OH, Zhu P, Ostendorff HP, et al., 2003. The histone deacetylase inhibitor valproic acid selectively induces proteasomal degradation of HDAC2. EMBO J, 22(13): 3411–3420. https://doi.org/10.1093/emboj/cdg315
Matsuzaki J, Tsuji T, Luescher IF, et al., 2015. Direct tumor recognition by a human CD4+ T-cell subset potently mediates tumor growth inhibition and orchestrates antitumor immune responses. Sci Rep, 5:14896. https://doi.org/10.1038/srep14896
Mojic M, Takeda K, Hayakawa Y, 2018. The dark side of IFN-γ: its role in promoting cancer immunoeevasion. Int J Mol Sci, 19(1):89. https://doi.org/10.3390/ijms19010089
Montgomery RL, Davis CA, Potthoff MJ, et al., 2007. Histone deacetylases 1 and 2 redundantly regulate cardiac morphogenesis, growth, and contractility. Gene Dev, 21(14): 1790–1802. https://doi.org/10.1101/gad.1563807
Müller BM, Jana L, Kasajima A, et al., 2013. Differential expression of histone deacetylases HDAC1, 2 and 3 in human breast cancer-overexpression of HDAC2 and HDAC3 is associated with clinicopathological indicators of disease progression. BMC Cancer, 13:215. https://doi.org/10.1186/1471-2407-13-21.
Muss HB, Caponera M, Zekan PJ, et al., 1986. Recombinant gamma interferon in advanced breast cancer: a phase II trial. Invest New Drugs, 4(4):377–381. https://doi.org/10.1007/BF00173511
Nam SW, Park JY, Ramasamy A, et al., 2005. Molecular changes from dysplastic nodule to hepatocellular carcinoma through gene expression profiling. Hepatology, 42(4): 809–818. https://doi.org/10.1002/hep.20878
Noh JH, Jung KH, Kim JK, et al., 2011. Aberrant regulation of HDAC2 mediates proliferation of hepatocellular carcinoma cells by deregulating expression of G1/S cell cycle proteins. PLoS ONE, 6(11):e28103. https://doi.org/10.1371/journal.pone.0028103
Ren WB, Li Y, Xia XJ, et al., 2018. Arginine inhibits the malignant transformation induced by interferon-gamma through the NF-κB-GCN2/eIF2α signaling pathway in mammary epithelial cells in vitro and in vivo. Exp Cell Res, 368(2):236–247. https://doi.org/10.1016/j.yexcr.2018.05.003
Sarhan D, D'Arcy P, Wennerberg E, et al., 2013. Activated monocytes augment TRAIL-mediated cytotoxicity by human NK cells through release of IFN-γ. Euro J Immunol, 43(1):249–257. https://doi.org/10.1002/eji.201242735
Schiller JH, Pugh M, Kirkwood JM, et al., 1996. Eastern cooperative group trial of interferon gamma in metastatic melanoma: an innovative study design. Clin Cancer Res, 2(1):29–36.
Trivedi CM, Zhu WT, Wang QH, et al., 2010. Hopx and Hdac2 interact to modulate Gata4 acetylation and embryonic cardiac Myocyte proliferation. Dev Cell, 19(3):450–459. https://doi.org/10.1016/j.devcel.2010.08.012
Wang JYJ, 2014. The capable ABL: what is its biological function? Mol Cell Biol, 34(7):1188–1197. https://doi.org/10.1128/MCB.01454-1.
Weichert W, Röske A, Gekeler V, et al., 2008. Histone deacetylases 1, 2 and 3 are highly expressed in prostate cancer and HDAC2 expression is associated with shorter PSA relapse time after radical prostatectomy. Br J Cancer, 98(3):604–610. https://doi.org/10.1038/sj.bjc.6604199
Xia XJ, Che YY, Gao YY, et al., 2016a. Arginine supplementation recovered the IFN-γ-mediated decrease in milk protein and fat synthesis by inhibiting the GCN2/eIF2α pathway, which induces autophagy in primary bovine mammary epithelial cells. Mol Cell, 39(5):410–417. https://doi.org/10.14348/molcells.2016.2358
Xia XJ, Gao YY, Zhang J, et al., 2016b. Autophagy mediated by arginine depletion activation of the nutrient sensor GCN2 contributes to interferon-γ-induced malignant transformation of primary bovine mammary epithelial cells. Cell Death Discov, 2:15065. https://doi.org/10.1038/cddiscovery.2015.65
Xia XJ, Che YY, Zhang J, et al., 2016c. Diet-driven interferon-γ enhances malignant transformation of primary bovine mammary epithelial cells through nutrient sensor GCN2-activated autophagy. Cell Death Dis, 7(3):e2138. https://doi.org/10.1038/cddis.2016.48
Yamaguchi T, Cubizolles F, Zhang Y, et al., 2010. Histone deacetylases 1 and 2 act in concert to promote the G1-to-S progression. Genes Dev, 24(5):455–469. https://doi.org/10.1101/gad.552310
Zaidi MR, Merlino G, 2011. The two faces of interferon-γ in cancer. Clin Cancer Res, 17(19):6118–6124. https://doi.org/10.1158/1078-0432.CCR-11-048.
Zhao HJ, Ho PC, Lo YH, et al., 2012. Interaction of proliferation cell nuclear antigen (PCNA) with c-Abl in cell proliferation and response to DNA damages in breast cancer. PLoS ONE, 7(1):e29416. https://doi.org/10.1371/journal.pone.0029416
Zhao HJ, Chen MS, Lo YH, et al., 2013. The Ron receptor tyrosine kinase activates c-Abl to promote cell proliferation through tyrosine phosphorylation of PCNA in breast cancer. Oncogene, 33(11):1429–1437. https://doi.org/10.1038/onc.2013.84
Zhou HY, Cai Y, Liu DN, et al., 2018. Pharmacological or transcriptional inhibition of both HDAC1 and 2 leads to cell cycle blockage and apoptosis via p21Waf1/Cip1 and p19INK4d upregulation in hepatocellular carcinoma. Cell Proliferat, 51(3):e12447. https://doi.org/10.1111/cpr.12447
Zhu JF, Yamane H, Paul WE, 2010. Differentiation of effector CD4 T cell populations. Annu Rev Immunol, 28:445–489. https://doi.org/10.1146/annurev-immunol-030409-10121.
Zuo H, Tell GS, Vollset SE, et al., 2014. Interferon-γ-induced inflammatory markers and the risk of cancer: the hordaland health study. Cancer, 120(21):3370–3377. https://doi.org/10.1002/cncr.28869
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Project supported by the National Natural Science Foundation of China (No. 31772715)
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Ren, Wb., Xia, Xj., Huang, J. et al. Interferon-γ regulates cell malignant growth via the c-Abl/HDAC2 signaling pathway in mammary epithelial cells. J. Zhejiang Univ. Sci. B 20, 39–48 (2019). https://doi.org/10.1631/jzus.B1800211
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DOI: https://doi.org/10.1631/jzus.B1800211