Aberrant expression of embryonic epithelial-mesenchymal transition-inducing transcription factors (EMT-TFs) in epithelial cells triggers EMT, neoplastic transformation, stemness, and metastatic dissemination. We found that regulation and functions of EMT-TFs are different in malignant melanoma. SNAIL2 and ZEB2 transcription factors are expressed in normal melanocytes and behave as tumor-suppressor proteins by activating an MITF-dependent melanocyte differentiation program. In response to NRAS/BRAF activation, EMT-TF network undergoes a profound reorganization in favor of TWIST1 and ZEB1. This reversible switch cooperates with BRAF in promoting dedifferentiation and neoplastic transformation of melanocytes. We detected EMT-TF reprogramming in late-stage melanoma in association with enhanced phospho-ERK levels. This switch results in E-cadherin loss, enhanced invasion, and constitutes an independent factor of poor prognosis in melanoma patients.

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Embryonic differentiation programs of epithelial-mesenchymal and mesenchymal-epithelial transition (EMT and MET) represent a mechanistic basis for epithelial cell plasticity implicated in cancer. Transcription factors of the ZEB protein family (ZEB1 and ZEB2) and several microRNA species (predominantly miR-200 family members) form a double negative feedback loop, which controls EMT and MET programs in both development and tumorigenesis. In this article, we review crosstalk between the ZEB/miR-200 axis and several signal transduction pathways activated at different stages of tumor development. The close association of ZEB proteins with these pathways is indirect evidence for the involvement of a ZEB/miR-200 loop in tumor initiation, progression and spread. Additionally, the configuration of signaling pathways involving ZEB/miR-200 loop suggests that ZEB1 and ZEB2 may have different, possibly even opposing, roles in some forms of human cancer.

The involvement of Mts1(S100A4), a small Ca 2+ -binding protein in tumor progression and metastasis had been demonstrated. However, the mechanism by which mts1(S100A4) promoted metastasis had not been identi®ed. Here we demonstrated that Mts1(S100A4) had signi®cant stimulatory eect on the angiogenesis. We detected high incidence of hemangiomas ± benign tumors of vascular origin in aged transgenic mice ubiquitously expressing the mts1(S100A4) gene. Furthermore, the serum level of the Mts1(S100A4) protein increased with ageing. Tumors developed in Mts1-transgenic mice revealed an enhanced vascular density. We showed that an oligomeric, but not a dimeric form of the Mts1(S100A4) protein was capable of enhancing the endothelial cell motility in vitro and stimulate the corneal neovascularization in vivo. An oligomeric fraction of the protein was detected in the conditioned media as well as in human serum. The data obtained allowed us to conclude that mts1(S100A4) might induce tumor progression via stimulation of angiogenesis. Oncogene (2001) 20, 4685 ± 4695.

S100A4(mts1) protein expression has been strongly associated with metastatic tumor progression. It has been suggested as a prognostic marker for a number of human cancers. It is proposed that extracellular S100A4 accelerates cancer progression by stimulating the motility of endothelial cells, thereby promoting angiogenesis. Here we show that in 3D culture mouse endothelial cells (SVEC 4-10) respond to recombinant S100A4 by stimulating invasive growth of capillary-like structures. The outgrowth is not dependent on the stimulation of cell proliferation, but rather correlates with the transcriptional modulation of genes involved in the proteolytic degradation of extracellular matrix (ECM). Treatment of SVEC 4-10 with the S100A4 protein leads to the transcriptional activation of collagenase 3 (MMP-13) mRNA followed by subsequent release of the protein from the cells. b-Casein zymography demonstrates enhancement of proteolytic activity associated with MMP-13. This observation indicates that extracellular S100A4 stimulates the production of ECM degrading enzymes from endothelial cells, thereby stimulating the remodeling of ECM. This could explain the angiogenic and metastasis-stimulating activity of S100A4(mts1).

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...

Zinc finger transcription factors of the Snail/Slug and ZEB-1/SIP1 families control epithelial-mesenchymal transitions in development in cancer. Here, we studied SIP1-regulated mesenchymal conversion of epidermoid A431 cells. We found that concomitant with inducing invasive phenotype, SIP1 inhibited expression of cyclin D1 and induced hypophosphorylation of the Rb tumor suppressor protein. Repression of cyclin D1 was caused by direct binding of SIP1 to three sequence elements in the cyclin D1 gene promoter. By expressing exogenous cyclin D1 in A431/SIP1 cells and using RNA interference, we demonstrated that the repression of cyclin D1 gene by SIP1 was necessary and sufficient for Rb hypophosphorylation and accumulation of cells in G1 phase. A431 cells expressing SIP1 along with exogenous cyclin D1 were highly invasive, indicating that SIP1-regulated invasion is independent of attenuation of G1/S progression. However, in another epithelial-mesenchymal transition model, gradual mesenchymal conversion of A431 cells induced by a dominant negative mutant of E-cadherin produced no effect on the cell cycle. We suggest that impaired G1/S phase progression is a general feature of cells that have undergone EMT induced by transcription factors of the Snail/Slug and ZEB-1/SIP1 families.

Two cell lines originating from a common ancestral tumor, CSML0 and CSML100, were used as a model to study AP-1 transcription factors at different steps of tumor progression. CSML0 cells have an epithelial morphology; they express epithelial but not mesenchymal markers and are invasive neither in vitro nor in vivo. CSML100 possesses all characteristics of a highly progressive carcinoma. These cells do not form tight contacts, are highly invasive in vitro, and are metastatic in vivo. AP-1 activity was considerably higher in CSML100 cells than in CSML0 cells. There was a common predominant Jun component, namely, JunD, detected in both cell lines. We found that the enhanced level of AP-1 in CSML100 cells was due to high expression of Fra-1 and Fra-2 proteins, which were undetectable in CSML0 nuclear extracts. Analysis of the transcription of different AP-1 members in various cell lines derived from tumors of epithelial origin revealed a correlation of fra-1 expression with mesenchymal characteristics of carcinoma cells. Moreover, we show here for the first time that the expression of exogenous Fra-1 in epithelioid cells results in morphological changes that resemble fibroblastoid conversion. Cells acquire an elongated shape and become more motile and invasive in vitro. Morphological alterations were accompanied by transcriptional activation of certain genes whose expression is often induced at late stages of tumor progression. These data suggest a critical role of the Fra-1 protein in the development of epithelial tumors.