Multistep carcinogenesis involves more than six discrete events also important in normal development and cell behavior. Of these, local invasion and metastasis cause most cancer deaths but are the least well understood molecularly. We employed a combined in vitro/in vivo carcinogenesis model, that is, polarized Ha-Ras–transformed mammary epithelial cells (EpRas), to dissect the role of Ras downstream signaling pathways in epithelial cell plasticity, tumorigenesis, and metastasis. Ha-Ras cooperates with transforming growth factor β (TGFβ) to cause epithelial mesenchymal transition (EMT) characterized by spindle-like cell morphology, loss of epithelial markers, and induction of mesenchymal markers. EMT requires continuous TGFβ receptor (TGFβ-R) and oncogenic Ras signaling and is stabilized by autocrine TGFβ production. In contrast, fibroblast growth factors, hepatocyte growth factor/scatter factor, or TGFβ alone induce scattering, a spindle-like cell phenotype fully reversible after factor withdrawal, which does not involve sustained marker changes. Using specific inhibitors and effector-specific Ras mutants, we show that a hyperactive Raf/mitogen-activated protein kinase (MAPK) is required for EMT, whereas activation of phosphatidylinositol 3-kinase (PI3K) causes scattering and protects from TGFβ-induced apoptosis. Hyperactivation of the PI3K pathway or the Raf/MAPK pathway are sufficient for tumorigenesis, whereas EMT in vivo and metastasis required a hyperactive Raf/MAPK pathway. Thus, EMT seems to be a close in vitro correlate of metastasis, both requiring synergism between TGFβ-R and Raf/MAPK signaling.

In contrast to the aberrant control of proliferation, apoptosis, angiogenesis and lifespan, the cellular mechanisms that cause local invasion and metastasis of tumour cells are still poorly understood. New experimental approaches have identified different types of epithelial-plasticity changes in tumour cells towards fibroblastoid phenotypes as crucial events that occur during metastasis, and many molecules and signalling pathways cooperate to trigger these processes.

Extensive prior research has focused on somatic copy-number alterations (SCNAs) affecting cancer genes, yet the extent to which recurrent SCNAs exert their influence through rearranging cis-regulatory elements remains unclear. Here, we present a framework for inferring cancer-related gene overexpression resulting from cis-regulatory element reorganization (e.g., enhancer hijacking), by integrating SCNAs, gene expression data, and information on chromatin interaction domains. Analysis of 7,416 cancer genomes uncovered several pan-cancer candidate genes, including IRS4, SMARCA1 and TERT. We demonstrate that IRS4 overexpression in lung cancer associates with recurrent deletions in cis, and present evidence supporting a tumor-promoting role. We additionally pursued cancer type-specific analyses, uncovering IGF2 as a target for enhancer hijacking in colorectal cancer. IGF2-containing tandem duplications result in the de novo formation of a 3D contact domain comprising IGF2 and a lineage-specific super-enhancer, which mediates high-level gene activation. Our framework enables systematic inference of cis-regulatory element rearrangements mediating dysregulation in cancer.

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Erk/MAPK and TGFbeta signaling cause epithelial to mesenchymal transition (EMT) and metastasis in mouse mammary epithelial cells (EpH4) transformed with oncogenic Ras (EpRas). In trials to unravel underlying mechanisms, expression profiling for EMT-specific genes identified a secreted interleukin-related protein (ILEI), upregulated exclusively at the translational level. Stable overexpression of ILEI in EpH4 and EpRas cells caused EMT, tumor growth, and metastasis, independent of TGFbeta-R signaling and enhanced by Bcl2. RNAi-mediated knockdown of ILEI in EpRas cells before and after EMT (EpRasXT) prevented and reverted TGFbeta-dependent EMT, also abrogating metastasis formation. ILEI is overexpressed and/or altered in intracellular localization in multiple human tumors, an event strongly correlated to invasion/EMT, metastasis formation, and survival in human colon and breast cancer.