Reprogramming of replicative senescence in hepatocellular carcinoma-derived cells

Öztürk, Nuri; Erdal, Esra; Mumcuoglu, Mine; Akçalı, Kamil Can; Yalçın, Özden; Şentürk, Şerif; Arslan-Ergül, Ayça; Gur, B; Yuluğ, Işık G.; Cetin-Atalay, Rengül; Yakıcıer, Cengiz; Yağcı, Tamer; Tez, Mesut; Öztürk, Mehmet

doi:10.1073/pnas.0510877103

Cited by 53 publications

(65 citation statements)

References 33 publications

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“…Inducible expression systems were used in most of the previous studies involving ZEB-2 overexpression (Comijn et al, 2001;Vandewalle et al, 2005), which is difficult to adapt for MCF10A cells. ZEB-2 has recently been shown to induce replicative senescence in certain cell types, suggesting that the activity of ZEB-2 is dependent on gene dose as well as cellular context (Ozturk et al, 2006). We observed the loss or reduced expression of p63 in both p65 and ZEB-1 overexpressing cells.…”

Section: Discussionsupporting

confidence: 48%

NF-κB represses E-cadherin expression and enhances epithelial to mesenchymal transition of mammary epithelial cells: potential involvement of ZEB-1 and ZEB-2

et al. 2006

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The transcription factor nuclear factor kappa B (NF-jB) is constitutively active in both cancer cells and stromal cells of breast cancer; however, the precise role of activated NF-jB in cancer progression is not known. Using parental MCF10A cells and a variant that expresses the myoepithelial marker p63 stably overexpressing the constitutively active p65 subunit of NFjB (MCF10A/p65), we show that NF-jB suppresses the expression of epithelial specific genes E-cadherin and desmoplakin and induces the expression of the mesenchymal specific gene vimentin. P65 also suppressed the expression of p63 and the putative breast epithelial progenitor marker cytokeratin 5/6. MCF10A/p65 cells were phenotypically similar to cells undergoing epithelial to mesenchymal transition (EMT). MCF10A/p65 cells failed to form characteristic acini in three-dimensional Matrigel. Analysis of parental and MCF10A/p65 cells for genes previously shown to be involved in EMT revealed elevated expression of ZEB-1 and ZEB-2 in MCF10A/ p65 cells compared to parental cells. In transient transfection assays, p65 increased ZEB-1 promoter activity. Furthermore, MCF10A cells overexpressing ZEB-1 showed reduced E-cadherin and p63 expression and displayed an EMT phenotype. The siRNA against ZEB-1 or ZEB-2 reduced the number of viable MCF10A/ p65 but not parental cells, suggesting the dependence of MCF10A/p65 cells to ZEB-1 and ZEB-2 for cell cycle progression or survival. MCF10A cells chronically exposed to tumor necrosis factor alpha (TNFa), a potent NF-jB inducer, also exhibited the EMT-like phenotype and ZEB-1/ZEB-2 induction, both of which were reversed following TNFa withdrawal.

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Section: Discussionsupporting

confidence: 48%

NF-κB represses E-cadherin expression and enhances epithelial to mesenchymal transition of mammary epithelial cells: potential involvement of ZEB-1 and ZEB-2

et al. 2006

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“…In fact, SIP1 was shown to directly repress cyclin D1 (Mejlvang et al, 2007). Also, induced expression of SIP1 was reported to be partly responsible for hTERT repression in hepatocellular carcinoma cells (Ozturk et al, 2006). On the other hand, SIP1 may be implicated in tumor development irrespective of its role in inducing EMT.…”

Section: Discussionmentioning

confidence: 99%

“…hTERT repression in breast cancer cells was partly mediated by SIP1 in a TGF-β dependent manner (Lin and Elledge, 2003). Also, analysis of senescence arrest of clonal hepatocellular carcinoma cells revealed SIP1 as a mediator of hTERT repression (Ozturk et al, 2006). Impaired G1/S progression was observed upon repression of cyclin D1 by SIP1 (Mejlvang et al, 2007).…”

Section: Introductionmentioning

confidence: 96%

Novel monoclonal antibodies detect Smad-interacting protein 1 (SIP1) in the cytoplasm of human cells from multiple tumor tissue arrays

Öztaş

Avci

Özcan

et al. 2010

Experimental and Molecular Pathology

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Smad-interacting protein 1 (SIP1, also known as ZEB2) represses the transcription of E-cadherin and mediates epithelial-mesenchymal transition in development and tumor metastasis. Due to the lack of human SIP1-specific antibodies, its expression in human tumor tissues has not been studied in detail by immunohistochemistry. Hence, we generated two anti-SIP1 monoclonal antibodies, clones 1C6 and 6E5, with IgG1 and IgG2a isotypes, respectively. The specificity of these antibodies was shown by Western blotting studies using siRNA mediated downregulation of SIP1 and ZEB1 in a human osteosarcoma cell line. In the same context, we also compared them with 5 commercially available SIP1 antibodies. Antibody specificity was further verified in an inducible cell line system by immunofluorescence. By using both antibodies, we evaluated the tissue expression of SIP1 in paraffin-embedded tissue microarrays consisting of 22 normal and 101 tumoral tissues of kidney, colon, stomach, lung, esophagus, uterus, rectum, breast and liver. Interestingly, SIP1 predominantly displayed a cytoplasmic expression, while the nuclear localization of SIP1 was observed in only 6 cases. Strong expression of SIP1 was found in distal tubules of kidney, glandular epithelial cells of stomach and hepatocytes, implicating a co-expression of SIP1 and E-cadherin. Squamous epithelium of the esophagus and surface epithelium of colon and rectum were stained with moderate to weak intensity. Normal uterus, breast and lung tissues remained completely negative. By comparison with their normal tissues, we observed SIP1 overexpression in cancers of the kidney, breast, lung and uterus. However, SIP1 expression was found to be downregulated in tumors from colon, rectum, esophagus, liver and stomach tissues. Finally we did nuclear/cytoplasmic fractionation in 3 carcinoma cell lines and detected SIP1 in both fractions, nucleus being the dominant one. To our best knowledge, this is the first comprehensive immunohistochemical study of the expression of SIP1 in a series of human cancers. Our finding that SIP1 is not exclusively localized to nucleus suggests that the subcellular localization of SIP1 is regulated in normal and tumor tissues. These novel monoclonal antibodies may help elucidate the role of SIP1 in tumor development.

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“…Here we demonstrated that SIP1 significantly repressed G 1 /S transition in RT112 cells, whereas the effect of ZEB1 was insignificant. In hepatocellular carcinoma cells, SIP1 repressed hTERT, resulting in replicative senescence (24). In contrast, ZEB1 was required for proliferation of mouse embryo fibroblasts, and ZEB1 Ϫ/Ϫ mouse embryo fibroblasts underwent premature replicative senescence (25).…”

Section: Discussionmentioning

confidence: 99%

SIP1 protein protects cells from DNA damage-induced apoptosis and has independent prognostic value in bladder cancer

Sayan

Griffiths

Pal

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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172

163

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The epithelial-mesenchymal transition (EMT) contributes to cancer metastasis. Two ZEB family members, ZEB1 and ZEB2(SIP1), inhibit transcription of the E-cadherin gene and induce EMT in vitro. However, their relevance to human cancer is insufficiently studied. Here, we performed a comparative study of SIP1 and ZEB1 proteins in cancer cell lines and in one form of human malignancy, carcinoma of the bladder. Whereas ZEB1 protein was expressed in all E-cadherin-negative carcinoma cell lines, being in part responsible for the high motility of bladder cancer cells, SIP1 was hardly ever detectable in carcinoma cells in culture. However, SIP1 represented an independent factor of poor prognosis (P ‫؍‬ 0.005) in a series of bladder cancer specimens obtained from patients treated with radiotherapy. In contrast, ZEB1 was rarely expressed in tumor tissues; and E-cadherin status did not correlate with the patients' survival. SIP1 protected cells from UV-and cisplatin-induced apoptosis in vitro but had no effect on the level of DNA damage. The anti-apoptotic effect of SIP1 was independent of either cell cycle arrest or loss of cell-cell adhesion and was associated with reduced phosphorylation of ATM/ATR targets in UV-treated cells. The prognostic value of SIP1 and its role in DNA damage response establish a link between genetic instability and metastasis and suggest a potential importance for this protein as a therapeutic target. In addition, we conclude that the nature of an EMT pathway rather than the deregulation of E-cadherin per se is critical for the progression of the disease and patients' survival. E pithelial mesenchymal transition (EMT) is a genetic program controlling cell migration during embryonic development and in wound healing (1, 2). Aberrant activation of EMT programs occurs in cells of epithelial tumors and contributes to the formation of cancer stem cells and metastasis (1-4). EMT is characterized by the loss of epithelial and the acquisition of mesenchymal features. EMT programs are controlled by several master regulators including TWIST, SNAIL (SNAI1 and SNAI2), and ZEB (ZEB1/␦EF1/TCF8 and SIP1/ZEB2) protein family members. These proteins act downstream in EMTinducing signal transduction pathways activated by growth factors, integrin engagement and hypoxia (1-3). Their expression is tightly regulated at the posttranscriptional level. Recent reports highlighted the importance of miR-200 microRNA family in the regulation of ZEB1 and SIP1 protein expression (5). ZEB proteins bind proximal E-boxes within the E-cadherin gene (cdh1) promoter and repress transcription by recruiting corepressor complexes (6). Likewise, they directly repress numerous genes encoding components of the epithelial junctional complex and cell polarity factors (7,8). The relevance of ZEB proteins to tumor progression has been studied in several forms of human cancer. Expression of ZEB1 correlated with the aggressive phenotype in various histological types of endometrial carcinoma and was detected in sarcomatous compartment of endometri...

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Reprogramming of replicative senescence in hepatocellular carcinoma-derived cells

Abstract: immortality ͉ liver cancer ͉ SIP1 ͉ telomerase ͉ p53

Cited by 53 publications

References 33 publications

NF-κB represses E-cadherin expression and enhances epithelial to mesenchymal transition of mammary epithelial cells: potential involvement of ZEB-1 and ZEB-2

NF-κB represses E-cadherin expression and enhances epithelial to mesenchymal transition of mammary epithelial cells: potential involvement of ZEB-1 and ZEB-2

Novel monoclonal antibodies detect Smad-interacting protein 1 (SIP1) in the cytoplasm of human cells from multiple tumor tissue arrays

SIP1 protein protects cells from DNA damage-induced apoptosis and has independent prognostic value in bladder cancer

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