Ets-1 regulates energy metabolism in cancer cells.

Cancer cells predominantly utilize glycolysis for ATP production even in the presence of abundant oxygen, an environment that would normally result in energy production through oxidative phosphorylation. Although the molecular mechanism for this metabolic switch to aerobic glycolysis has not been fully elucidated, it is likely that mitochondrial damage to the electron transport chain and the resulting increased production of reactive oxygen species are significant driving forces. In this study, we have investigated the role of the transcription factor Ets-1 in the regulation of mitochondrial function and metabolism. Ets-1 was over-expressed using a stably-incorporated tetracycline-inducible expression vector in the ovarian cancer cell line 2008, which does not express detectable basal levels of Ets-1 protein. Microarray analysis of the effects of Ets-1 over-expression in these ovarian cancer cells shows that Ets-1 up-regulates key enzymes involved in glycolysis and associated feeder pathways, fatty acid metabolism, and antioxidant defense. In contrast, Ets-1 down-regulates genes involved in the citric acid cycle, electron transport chain, and mitochondrial proteins. At the functional level, we have found that Ets-1 expression is directly correlated with cellular oxygen consumption whereby increased expression causes decreased oxygen consumption. Ets-1 over-expression also caused increased sensitivity to glycolytic inhibitors, as well as growth inhibition in a glucose-depleted culture environment. Collectively our findings demonstrate that Ets-1 is involved in the regulation of cellular metabolism and response to oxidative stress in ovarian cancer cells.

Mechanisms associated with mitochondrial-generated reactive oxygen species in cancer.

The mitochondria are unique cellular organelles that contain their own genome and, in conjunction with the nucleus, are able to transcribe and translate genes encoding components of the electron transport chain (ETC). To do so, the mitochondria must communicate with the nucleus via the production of reactive oxygen species (ROS) such as hydrogen peroxide (H2O2), which are produced as a byproduct of aerobic respiration within the mitochondria. Mitochondrial signaling is proposed to be altered in cancer cells, where the mitochondria are frequently found to harbor mutations within their genome and display altered functional characteristics leading to increased glycolysis. As signaling molecules, ROS oxidize and inhibit MAPK phosphatases resulting in enhanced proliferation and survival, an effect particularly advantageous to cancer cells. In terms of transcriptional regulation, ROS affect the phosphorylation, activation, oxidation, and DNA binding of transcription factors such as AP-1, NF-kappaB, p53, and HIF-1alpha, leading to changes in target gene expression. Increased ROS production by defective cancer cell mitochondria also results in the upregulation of the transcription factor Ets-1, a factor that has been increasingly associated with aggressive cancers.

VHL promotes E2 box-dependent E-cadherin transcription by HIF-mediated regulation of SIP1 and snail.

The product of the von Hippel-Lindau gene (VHL) acts as the substrate-recognition component of an E3 ubiquitin ligase complex that ubiquitylates the catalytic alpha subunit of hypoxia-inducible factor (HIF) for oxygen-dependent destruction. Although emerging evidence supports the notion that deregulated accumulation of HIF upon the loss of VHL is crucial for the development of clear-cell renal cell carcinoma (CC-RCC), the molecular events downstream of HIF governing renal oncogenesis remain unclear. Here, we show that the expression of a homophilic adhesion molecule, E-cadherin, a major constituent of epithelial cell junctions whose loss is associated with the progression of epithelial cancers, is significantly down-regulated in primary CC-RCC and CC-RCC cell lines devoid of VHL. Reintroduction of wild-type VHL in CC-RCC (VHL(-/-)) cells markedly reduced the expression of E2 box-dependent E-cadherin-specific transcriptional repressors Snail and SIP1 and concomitantly restored E-cadherin expression. RNA interference-mediated knockdown of HIFalpha in CC-RCC (VHL(-/-)) cells likewise increased E-cadherin expression, while functional hypoxia or expression of VHL mutants incapable of promoting HIFalpha degradation attenuated E-cadherin expression, correlating with the disengagement of RNA polymerase II from the endogenous E-cadherin promoter/gene. These findings reveal a critical HIF-dependent molecular pathway connecting VHL, an established "gatekeeper" of the renal epithelium, with a major epithelial tumor suppressor, E-cadherin.

ets-1 is transcriptionally up-regulated by H2O2 via an antioxidant response element.

Expression of the transcription factor Ets-1 is increasingly associated with the progression of several human cancers. A tumor-derived factor is expected to be involved in the inappropriate up-regulation of ets-1 in tumor and surrounding cells. A link between hydrogen peroxide (H2O2) and increased Ets-1 expression has also been suggested, leading to the proposal that this reactive oxygen species (ROS) may be an important factor in directly regulating the expression of ets-1 in tumor cells. Ets-1 expression in response to H2O2 was examined in an ovarian carcinoma cell model, and the genes promoter region was analyzed in order to identify putative elements involved in redox responsiveness. The up-regulation of Ets-1 by H2O2 was confirmed in the cells tested. Luciferase assays using constructs generated to test the contribution of specific promoter elements indicated that an antioxidant response element (ARE) is primarily involved in the H2O2-mediated induction. Gel shift analysis confirmed the increased binding of an Nrf2 containing protein complex to the ets-1 ARE after H2O2 treatment. This study has delineated a key element involved in the transcriptional regulation of ets-1 under basal and induced conditions. Ets-1 has obvious deleterious effects in many cancer cells, and thus, the identification of this regulatory pathway has provided possible targets for manipulating its expression.

Role of the transcription factor Ets-1 in cisplatin resistance.

Cisplatin is a DNA damaging agent widely used as a chemotherapeutic agent. A major limitation of the use of this agent is the development of drug resistance within tumors. Several in vitro models exist which enable the investigation of resistance mechanisms, including 2008/C13* ovarian carcinoma cells. C13* cells are variants of 2008 cells, displaying cisplatin resistance following 13 consecutive cisplatin treatments. This model system has led to the identification of several mechanisms that play parts in the multifactorial nature of cisplatin resistance. In this study, we have examined the contribution of a transcription factor, Ets-1, to the cisplatin resistance of C13* cells. Ets-1 is up-regulated in C13* cells as compared with the cisplatin-sensitive 2008 cells and overexpression of this protein in 2008 cells led to a 7-fold increase in resistance. Further studies on a colorectal carcinoma cell line overexpressing Ets-1 indicated that this phenomenon is not cell specific-increased cisplatin resistance correlated to Ets-1 expression. The mechanism of cisplatin resistance elicited by Ets-1 is potentially via transcriptional activation of genes whose products have well-described functions in reducing cisplatin toxicity. Examples, identified via microarray analysis, include metallothioneins and DNA repair enzymes. This is the first report to our knowledge associating expression of Ets-1, a transcription factor whose expression often signals poor prognosis in various cancer types, to cisplatin resistance.