Tian Yuan, Ding Wencheng, Wang Yingying, Ji Teng, Sun Shujuan, Mo Qingqing, Chen Pingbo, Fang Yong, Liu Jia, Wang Beibei, Zhou Jianfeng, Ma Ding, Wu Peng
Cancer Biology Research Center, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, P. R. of China.
Department of Gynecology, Third Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, P. R. of China.
PLoS One. 2013 Dec 18;8(12):e84457. doi: 10.1371/journal.pone.0084457. eCollection 2013.
Cervical cancer cells exhibit an increased requirement for ubiquitin-dependent protein degradation associated with an elevated metabolic turnover rate. Ubiquitin, which is a small, highly conserved protein expressed in all eukaryotic cells, can be covalently linked to certain target proteins to mark them for degradation by the ubiquitin-proteasome system. Previous studies highlight the essential role of Ubiquitin B (UbB) and UbB-dependent proteasomal protein degradation in histone deacetylase inhibitor (HDACi) -induced tumor selectivity. We hypothesized that UbB plays a critical role in the function of cervical cancer stem cells. We measured endogenous UbB levels in mammospheres in vitro by real-time PCR and Western blotting. The function of UbB in cancer stem-like cells was assessed after knockdown of UbB expression in prolonged Trichostatin A-selected HeLa cells (HeLa/TSA) by measuring in vitro cell proliferation, cell apoptosis, invasion, and chemotherapy resistance as well as by measuring in vivo growth in an orthotopic model of cervical cancer. We also assessed the cancer stem cell frequency, tumorsphere formation, and in vivo growth of human cervical cancer xenografts after UbB silencing. We found that HeLa/TSA were resistant to chemotherapy, highly expressed the UbB gene and the stem cell markers Sox2, Oct4 and Nanog. These cells also displayed induced differentiation abilities, including enhanced migration/invasion/malignancy capabilities in vitro and in vivo. Furthermore, an elevated expression of UbB was shown in the tumor samples of chemotherapy patients. Silencing of UbB inhibited tumorsphere formation, lowered the expression of stem cell markers and decreased cervical xenograft growth. Our results demonstrate that UbB was significantly increased in prolonged Trichostatin A-selected HeLa cells and it played a key role in the maintenance of cervical cancer stem-like cells.
宫颈癌细胞对泛素依赖性蛋白质降解的需求增加，这与代谢周转率升高有关。泛素是一种在所有真核细胞中表达的小的、高度保守的蛋白质，它可以与某些靶蛋白共价连接，以标记它们被泛素-蛋白酶体系统降解。先前的研究强调了泛素B（UbB）和UbB依赖性蛋白酶体蛋白降解在组蛋白去乙酰化酶抑制剂（HDACi）诱导的肿瘤选择性中的重要作用。我们假设UbB在宫颈癌干细胞的功能中起关键作用。我们通过实时PCR和蛋白质印迹法在体外测量了乳腺球中的内源性UbB水平。在通过测量体外细胞增殖、细胞凋亡、侵袭和化疗耐药性以及通过测量宫颈癌原位模型中的体内生长，在长期曲古抑菌素A选择的HeLa细胞（HeLa/TSA）中敲低UbB表达后，评估了UbB在癌症干细胞样细胞中的功能。我们还评估了UbB沉默后人宫颈癌异种移植瘤的癌症干细胞频率、肿瘤球形成和体内生长情况。我们发现HeLa/TSA对化疗耐药，高表达UbB基因以及干细胞标志物Sox2、Oct4和Nanog。这些细胞还表现出诱导分化能力，包括在体外和体内增强的迁移/侵袭/恶性能力。此外，化疗患者的肿瘤样本中显示UbB表达升高。UbB沉默抑制了肿瘤球形成，降低了干细胞标志物的表达，并减少了宫颈异种移植瘤的生长。我们的结果表明，在长期曲古抑菌素A选择的HeLa细胞中UbB显著增加，并且它在维持宫颈癌干细胞样细胞中起关键作用。
