Dissociation of EphB2 Signaling Pathways Mediating Progenitor Cell Proliferation and Tumor Suppression

SummarySignaling proteins driving the proliferation of stem and progenitor cells are often encoded by proto-oncogenes. EphB receptors represent a rare exception; they promote cell proliferation in the intestinal epithelium and function as tumor suppressors by controlling cell migration and inhibiting invasive growth. We show that cell migration and proliferation are controlled independently by the receptor EphB2. EphB2 regulated cell positioning is kinase-independent and mediated via phosphatidylinositol 3-kinase, whereas EphB2 tyrosine kinase activity regulates cell proliferation through an Abl-cyclin D1 pathway. Cyclin D1 regulation becomes uncoupled from EphB signaling during the progression from adenoma to colon carcinoma in humans, allowing continued proliferation with invasive growth. The dissociation of EphB2 signaling pathways enables the selective inhibition of the mitogenic effect without affecting the tumor suppressor function and identifies a pharmacological strategy to suppress adenoma growth

EphB Receptors Coordinate Migration and Proliferation in the Intestinal Stem Cell Niche

SummaryMore than 1010 cells are generated every day in the human intestine. Wnt proteins are key regulators of proliferation and are known endogenous mitogens for intestinal progenitor cells. The positioning of cells within the stem cell niche in the intestinal epithelium is controlled by B subclass ephrins through their interaction with EphB receptors. We report that EphB receptors, in addition to directing cell migration, regulate proliferation in the intestine. EphB signaling promotes cell-cycle reentry of progenitor cells and accounts for approximately 50% of the mitogenic activity in the adult mouse small intestine and colon. These data establish EphB receptors as key coordinators of migration and proliferation in the intestinal stem cell niche

Epicardial cell shape and maturation are regulated by Wt1 via transcriptional control of Bmp4

The epicardium plays a crucial role in embryonic heart development and adult heart repair; however, the molecular events underlying its maturation remain unknown. Wt1, one of the main markers of the embryonic epicardium, is essential for epicardial development and function. Here, we analyse the transcriptomic profile of epicardial-enriched cells at different stages of development and from control and epicardial-specific Wt1 knockout (Wt1KO) mice. Transcriptomic and cell morphology analyses of epicardial cells from epicardial-specific Wt1KO mice revealed a defect in the maturation process of the mutant epicardium, including sustained upregulation of Bmp4 expression and the inability of mutant epicardial cells to transition into a mature squamous phenotype. We identified Bmp4 as a transcriptional target of Wt1, thus providing a molecular basis for the retention of the cuboidal cell shape observed in the Wt1KO epicardium. Accordingly, inhibition of the Bmp4 signalling pathway both ex vivo and in vivo rescued the cuboidal phenotype of the mutant epicardium. Our findings indicate the importance of the cuboidal-to-squamous transition in epicardial maturation, a process regulated by Wt1

Ephrin-Bs Drive Junctional Downregulation and Actin Stress Fiber Disassembly to Enable Wound Re-epithelialization

For a skin wound to successfully heal, the cut epidermal-edge cells have to migrate forward at the interface between scab and healthy granulation tissue. Much is known about how lead-edge cells migrate, but very little is known about the mechanisms that enable active participation by cells further back. Here we show that ephrin-B1 and its receptor EphB2 are both upregulated in vivo, just for the duration of repair, in the first 70 or so rows of epidermal cells, and this signal leads to downregulation of the molecular components of adherens and tight (but not desmosomal) junctions, leading to loosening between neighbors and enabling shuffle room among epidermal cells. Additionally, this signaling leads to the shutdown of actomyosin stress fibers in these same epidermal cells, which may act to release tension within the wound monolayer. If this signaling axis is perturbed, then disrupted healing is a consequence in mouse and man. Epithelial migration is essential for skin wound healing

Enhancer decommissioning by Snail1-induced competitive displacement of TCF7L2 and down-regulation of transcriptional activators results in EPHB2 silencing

Transcriptional silencing is a major cause for the inactivation of tumor suppressor genes, however, the underlying mechanisms are only poorly understood. The EPHB2 gene encodes a receptor tyrosine kinase that controls epithelial cell migration and allocation in intestinal crypts. Through its ability to restrict cell spreading, EPHB2 functions as a tumor suppressor in colorectal cancer whose expression is frequently lost as tumors progress to the carcinoma stage. Previously we reported that EPHB2 expression depends on a transcriptional enhancer whose activity is diminished in EPHB2 non-expressing cells. Here we investigated the mechanisms that lead to EPHB2 enhancer inactivation. We show that expression of EPHB2 and SNAIL1 - an inducer of epithelial-mesenchymal transition (EMT) - is anti-correlated in colorectal cancer cell lines and tumors. In a cellular model of Snail1-induced EMT, we observe that features of active chromatin at the EPHB2 enhancer are diminished upon expression of murine Snail1. We identify the transcription factors FOXA1, MYB, CDX2 and TCF7L2 as EPHB2 enhancer factors and demonstrate that Snail1 indirectly inactivates the EPHB2 enhancer by downregulation of FOXA1 and MYB. In addition, Snail1 induces the expression of Lymphoid enhancer factor 1 (LEF1) which competitively displaces TCF7L2 from the EPHB2 enhancer. In contrast to TCF7L2, however, LEF1 appears to repress the EPHB2 enhancer. Our findings underscore the importance of transcriptional enhancers for gene regulation under physiological and pathological conditions and show that SNAIL1 employs a combinatorial mechanism to inactivate the EPHB2 enhancer based on activator deprivation and competitive displacement of transcription factors

Systemic Analysis of Gene Expression Profiles Identifies ErbB3 as a Potential Drug Target in Pediatric Alveolar Rhabdomyosarcoma

Peer reviewe

No Guts, no Glory : EphB mediated signaling in intestinal stem and progenitor cells

The work presented in this thesis is focused on the intestinal stem cell niche, the crypts, and the Eph tyrosine kinase receptors together with their ligands that are expressed by the crypt epithelial progenitor cells. Two fundamental cellular processes, high turnover coupled to a rapid ordered migration of the newborn cells out from the crypt, characterize the normal intestine. Transformed epithelial cells mirror these processes, gaining an advantage over the untransformed cells by the capacity of going through seemingly endless rounds of division, as well as expanding spatially into surrounding tissues by increased motility. The B class Eph receptors are involved in regulating both proliferation and migration in normal as well as transformed intestinal epithelial cells. In Paper I, we show that EphB receptors, in addition to directing cell migration, regulate proliferation in the intestine. We use both loss and gain of function experiments to correlate the EphB forward signaling to the rate of progenitor cell proliferation. EphB signaling promotes cell cycle reentry of progenitor cells and accounts for up to half the mitogenic activity in the adult intestine. We suggest a model where the restricted Wnt source plays an important role in establishing the EphBephrin- B gradients, hence extending the proliferative domain beyond the region directly influenced by the Wnt proteins themselves. In Paper II we continue to investigate the dual role of EphB receptors. We show that cell migration and proliferation are controlled independently by the EphB2 receptor. EphB2 regulated cell positioning is kinase-independent and mediated via PI3-kinase, whereas EphB2 tyrosine kinase activity regulates cell proliferation through an Abl-cyclin D1 pathway. Cyclin D1 regulation becomes uncoupled from EphB signaling during the progression from adenoma to colon carcinoma, allowing continued proliferation with invasive growth. The dissociation of EphB2 signaling pathways enables the selective inhibition of the mitogenic signaling pathway without affecting the tumor suppressor function of EphB2. In Paper III we investigate the role of the two ligands expressed in the intestinal epithelium, ephrin-B1 and ephrin-B2 and show that, although both ligands can bind EphB2 and EphB3, they have distinct functional outcomes when activating the receptors. Ephrin-B1 selectively affects migration, without influencing proliferation, whereas ephrin-B2 affects proliferation, but not migration

Generating Headlines with Recurrent Neural Networks

This report describes the implementation and evaluation of two natural language models using the machine learning technique deep learning. More specifically, two different models describing recurrent artificial neural networks (RNNs) were implemented, capable of generating news article headlines. One model focused on the generation of random (unconditioned) headlines, and the other one on the generation of headlines based (conditioned) on a given news article. Both models were then trained and evaluated on a data set of approximately 500,000 pairs of news articles and their corresponding headlines. The task of summarizing large bodies of text into smaller ones, while maintaining the key points of the original text, has many applications. Quickly and automatically obtaining condensed versions of for example medical journals, scientific papers, and news articles can be of great value for the users of such content. The unconditioned model, implemented using a multi-layer RNN consisting of LSTM cells, was able to produce headlines of moderate plausibility, a majority being syntactically correct. The conditioned model was implemented using two RNNs consisting of GRU cells in an encoder-decodernetwork with an attention mechanism, allowing the network to learn what words to focus on during headlining. Although the model managed to identify important keywords in the articles, it seldomly managed to produce meaningful sentences with them. We conclude that the techniques and models described in this report could be used to generate plausible news headlines. However, for the purpose of generating conditioned headlines, we think that additional modifications are needed to obtain satisfying results