The Role of Eph Receptors and Ephrins in Corneal Physiology and Diseases.

The cornea, while appearing to be simple tissue, is actually an extremely complex structure. In order for it to retain its biomechanical and optical properties, perfect organization of its cells is essential. Proper regeneration is especially important after injuries and in the course of various diseases. Eph receptors and ephrin are mainly responsible for the proper organization of tissues as well as cell migration and communication. In this review, we present the current state of knowledge on the role of Eph and ephrins in corneal physiology and diseases, in particular, we focused on the functions of the epithelium and endothelium. Since the role of Eph and ephrins in the angiogenesis process has been well established, we also analyzed their influence on conditions with corneal neovascularization.

Eph Receptors in the Immunosuppressive Tumor Microenvironment.

The tumor microenvironment (TME) promotes tumor development via complex intercellular signaling, aiding tumor growth and suppressing immunity. Eph receptors (Eph) and their ephrin ligands control cell interactions during normal development, and reemerge in tumors and the TME, where they are implicated in invasion, metastasis, and angiogenesis. Recent studies also indicate roles for Ephs in suppressing immune responses by controlling tumor interactions with innate and adaptive immune cells within the TME. Accordingly, inhibiting these functions can promote immune response and efficacy of immune checkpoint inhibition. This research highlights Ephs as potential targets to enhance efficacy of immune-based therapies in patients with cancer.

Role of forward and reverse signaling in Eph receptor and ephrin mediated cell segregation.

Eph receptor and ephrin signaling has a major role in segregating distinct cell populations to form sharp borders. Expression of interacting Ephs and ephrins typically occurs in complementary regions, such that polarised activation of both components occurs at the interface. Forward signaling through Eph receptors can drive cell segregation, but it is unclear whether reverse signaling through ephrins can also contribute. We have tested the role of reverse signaling, and of polarised versus non-polarised activation, in assays in which contact repulsion drives cell segregation and border sharpening. We find that polarised forward signaling drives stronger segregation than polarised reverse signaling. Nevertheless, reverse signaling contributes since bidirectional Eph and ephrin activation drives stronger segregation than unidirectional forward signaling alone. In contrast, non-polarised Eph activation drives little segregation. We propose that although polarised forward signaling is the principal driver of segregation, reverse signaling enables bidirectional repulsion which prevents mingling of each population into the other.

[Regulatory Effects of Erythropoietin-producing Hepatocyte Kinase Receptor and Ephrin Ligand on Bone Remodeling].

During the process of bone remodeling,the bone homeostasis is tightly controlled by the coupling of bone resorption and bone formation,depending upon cellular communication between osteoclasts and osteoblasts. Many studies have identified that the bi-directional transduction of erythropoietin producing hepatocyte kinase receptor and ephrin ligand (Eph/ephrin) is one of signal transduction pathways in bone remodeling. This review focus on the potential role of Eph/ephrin in bone remodeling,especially in alveolar remodeling.

Dying to communicate: apoptotic functions of Eph/Ephrin proteins.

The Erythropoietin-producing human hepatocellular carcinoma (Eph) receptors constitute the largest family of receptor tyrosine kinases and interact with a group of ligands called Ephrins. An essential feature of the Eph receptors and Ephrin ligands is that both are membrane-bound and, upon cell-cell interaction, initiate a bidirectional signaling involving both the receptor (forward signaling) and the ligand (reverse signaling). They regulate a large set of pleiotropic functions in virtually every tissue and physiological system. In vitro as well as in vivo data support a role for Eph and Ephrin molecules in cellular processes such as proliferation, cell-cell attraction and repulsion, motility and sorting. An increasing amount of evidence supports a role for these molecules in apoptosis and, although this function in cell death has been barely examined, the available information warrants a global consideration, to identify unmet needs and potential research avenues. Here we propose a comprehensive analysis of the data available regarding the importance of Ephs and Ephrins in cell death mechanisms throughout a large array of physiological systems.

The Wound Microenvironment Reprograms Schwann Cells to Invasive Mesenchymal-like Cells to Drive Peripheral Nerve Regeneration.

Schwann cell dedifferentiation from a myelinating to a progenitor-like cell underlies the remarkable ability of peripheral nerves to regenerate following injury. However, the molecular identity of the differentiated and dedifferentiated states in vivo has been elusive. Here, we profiled Schwann cells acutely purified from intact nerves and from the wound and distal regions of severed nerves. Our analysis reveals novel facets of the dedifferentiation response, including acquisition of mesenchymal traits and a Myc module. Furthermore, wound and distal dedifferentiated Schwann cells constitute different populations, with wound cells displaying increased mesenchymal character induced by localized TGFß signaling. TGFß promotes invasion and crosstalks with Eph signaling via N-cadherin to drive collective migration of the Schwann cells across the wound. Consistently, Tgfbr2 deletion in Schwann cells resulted in misdirected and delayed reinnervation. Thus, the wound microenvironment is a key determinant of Schwann cell identity, and it promotes nerve repair through integration of multiple concerted signals. VIDEO ABSTRACT.

α2-chimaerin is required for Eph receptor-class-specific spinal motor axon guidance and coordinate activation of antagonistic muscles.

Axonal guidance involves extrinsic molecular cues that bind growth cone receptors and signal to the cytoskeleton through divergent pathways. Some signaling intermediates are deployed downstream of molecularly distinct axon guidance receptor families, but the scope of this overlap is unclear, as is the impact of embryonic axon guidance fidelity on adult nervous system function. Here, we demonstrate that the Rho-GTPase-activating protein α2-chimaerin is specifically required for EphA and not EphB receptor signaling in mouse and chick spinal motor axons. Reflecting this specificity, the loss of α2-chimaerin function disrupts the limb trajectory of extensor-muscle-innervating motor axons the guidance of which depends on EphA signaling. These embryonic defects affect coordinated contraction of antagonistic flexor-extensor muscles in the adult, indicating that accurate embryonic motor axon guidance is critical for optimal neuromuscular function. Together, our observations provide the first functional evidence of an Eph receptor-class-specific intracellular signaling protein that is required for appropriate neuromuscular connectivity.

Therapeutic targeting of EPH receptors and their ligands.

Critical roles for EPH receptor (EPH)-ephrin signalling in a range of chronic and regenerative diseases are increasingly being recognized. In particular, the complex roles of EPHs and ephrins in tumour growth and progression, and in nerve injury and regeneration have been studied extensively. This has led to considerable progress in developing strategies for their therapeutic targeting, with some anticancer agents already in clinical trials. Promising leads for non-malignant diseases are also emerging, with compelling preclinical data encouraging clinical development. We discuss this rapidly developing area of drug discovery, highlighting the associated challenges and limitations.

Tie2 and Eph receptor tyrosine kinase activation and signaling.

The Eph and Tie cell surface receptors mediate a variety of signaling events during development and in the adult organism. As other receptor tyrosine kinases, they are activated on binding of extracellular ligands and their catalytic activity is tightly regulated on multiple levels. The Eph and Tie receptors display some unique characteristics, including the requirement of ligand-induced receptor clustering for efficient signaling. Interestingly, both Ephs and Ties can mediate different, even opposite, biological effects depending on the specific ligand eliciting the response and on the cellular context. Here we discuss the structural features of these receptors, their interactions with various ligands, as well as functional implications for downstream signaling initiation. The Eph/ephrin structures are already well reviewed and we only provide a brief overview on the initial binding events. We go into more detail discussing the Tie-angiopoietin structures and recognition.

Therapeutic perspectives of Eph-ephrin system modulation.

Eph receptors are the largest class of kinase receptors and, together with their ligands ephrins, they have a primary role in embryogenesis. Their expression has been found deregulated in several cancer tissues and, in many cases, abnormal levels of these proteins have been correlated to a poor prognosis. Recently, the Eph-ephrin system was found to be deregulated in other pathological processes, involving the nervous and cardiovascular systems. The increasing body of evidence supports the Eph-ephrin system as a target not only for the treatment of solid tumors, but also to face other critical diseases such as amyotrophic lateral sclerosis and diabetes driving current efforts toward the development of pharmacological tools potentially able to treat these pathologies.

The role of EPH receptors in cancer-related epithelial-mesenchymal transition.

Erythropoietin-producing hepatoma (EPH) receptors are considered the largest family of receptor tyrosine kinases and play key roles in physiological and pathologic processes in development and disease. EPH receptors are often overexpressed in human malignancies and are associated with poor prognosis. However, the functions of EPH receptors in epithelial-mesenchymal transition (EMT) remain largely unknown. This review depicts the relationship between EPH receptors and the EMT marker E-cadherin as well as the crosstalk between EPH receptors and the signaling pathways involved EMT. Further discussion is focused on the clinical significance of EPH receptors as candidates for targeting in cancer therapeutics. Finally, we summarize how targeted inhibition of both EPH receptors and EMT-related signaling pathways represents a novel strategy for cancer treatment.

Genetic analysis of left-right coordination of locomotion .

While there is a rather large amount of data from pharmacological and anatomical studies of the murine locomotor CPG network, comprehensive information regarding the cellular and functional properties of the neuronal populations is lacking. Here we describe concepts arising from genetic studies of the locomotor network with a focus on commissural interneurons regulating left-right coordination. In particular, this involves several families of axon guidance molecules relevant for midline crossing. We also describe recent advances within the field of neural circuit analysis, including imaging, genetic inactivation and optogenetic strategies, which are applicable to locomotor circuits. Such efforts, for example by using available genetic markers, should substantially increase our possibilities to decipher the functionality of spinal cord neuronal networks.

The ephrin signaling pathway regulates morphology and adhesion of mouse granulosa cells in vitro.

Follicle-stimulating hormone (FSH)-mediated changes in granulosa cell adhesion and morphology are essential for preovulatory follicle development, given the dramatic changes in follicle size and granulosa cell number that occur during this transition. Members of the Eph-ephrin family of cell-positioning and adhesion molecules, a family that consists of ephrin ligands and their Ephrin (Eph) receptors, regulate cell location, adhesion, and migration during embryonic development and tumor growth. However, very little is known about ephrin signaling during folliculogenesis. We have found that FSH increases the expression of several members of the Eph-ephrin family and that this signaling regulates granulosa cell morphology and adhesion. FSH induced increased mRNA levels of the ephrin ligand, ephrin-A5 (Efna5), and its receptors, Eph receptors A3, A5, and A8 (Epha3, Epha5, and Epha8, respectively), in granulosa cells. Immunofluorescence studies indicated that EFNA5 and EPHA5 are located in the membrane of granulosa cells of developing mouse follicles. Eph-ephrin signaling directly affected granulosa cell morphology and adhesion. Recombinant EFNA5 reduced cell spreading and increased cell rounding in mouse primary granulosa cells and in a rat granulosa cell line, whereas EPHA5 reduced granulosa cell adhesion in both model systems. Both FSH and forskolin also increased Efna5 and Epha5 mRNA levels in rat and human granulosa cell lines, indicating that FSH regulates these genes via the cAMP-dependent protein kinase A pathway and that this regulation is conserved across different species. The present study identifies Eph-ephrin signaling as a novel FSH-mediated pathway regulating granulosa cell morphology and adhesion.

Ephs and ephrins resurface in inflammation, immunity, and atherosclerosis.

Despite significant advancements in treatment regimens, cardiovascular disease remains a worldwide leader of morbidity, mortality, and healthcare cost. A large percentage of cardiovascular disease is directly attributable to the process of atherosclerosis, a chronic inflammatory disease of the vessel wall. In the hunt for novel therapeutic targets in cardiovascular disease, neuronal guidance molecules are emerging as significant regulators of cardiovascular remodeling and inflammation. The Eph family of neuronal guidance molecules comprises the largest family of receptor tyrosine kinases in the mammalian genome. While best characterized in embryogenesis and carcinogenesis, Eph receptors and their ephrin ligands are becoming increasingly recognized as important players in chronic inflammatory diseases and immune function. Herein we discuss the current evidence for how Eph/ephrin interactions, particularly EphA2/ephrinA1 and EphB/ephrinB2, affect inflammation and cardiovascular disease.

A chemical genetic approach reveals distinct EphB signaling mechanisms during brain development.

EphB receptor tyrosine kinases control multiple steps in nervous system development. However, it remains unclear whether EphBs regulate these different developmental processes directly or indirectly. In addition, given that EphBs signal through multiple mechanisms, it has been challenging to define which signaling functions of EphBs regulate particular developmental events. To address these issues, we engineered triple knock-in mice in which the kinase activity of three neuronally expressed EphBs can be rapidly, reversibly and specifically blocked. We found that the tyrosine kinase activity of EphBs was required for axon guidance in vivo. In contrast, EphB-mediated synaptogenesis occurred normally when the kinase activity of EphBs was inhibited, suggesting that EphBs mediate synapse development by an EphB tyrosine kinase-independent mechanism. Taken together, our data indicate that EphBs control axon guidance and synaptogenesis by distinct mechanisms and provide a new mouse model for dissecting EphB function in development and disease.

Signal transduction pathways participating in homeostasis and malignant transformation of the intestinal tissue.

Intestinal homeostasis is a complex and tightly regulated process governed by a variety of signalling pathways that balance cell proliferation and differentiation. As revealed by extensive use of defined mouse models, perturbations within the signalling circuitry trigger initial expansion of premalignant cells. In this review, we attempt to summarise recent advances in the knowledge of the cellular signalling mechanisms that drive tumorigenesis in the human and mouse intestine.

Ankyrin repeat-rich membrane spanning protein (kidins220) is required for neurotrophin and ephrin receptor-dependent dendrite development.

Dendrites are the primary sites on neurons for receiving and integrating inputs from their presynaptic partners. Defects in dendrite development perturb the formation of neural circuitry and impair information processing in the brain. Extracellular cues are important for shaping the dendritic morphogenesis, but the underlying molecular mechanisms are not well understood. In this study, we examined the role of ARMS (ankyrin repeat-rich membrane spanning protein), also known as Kidins220 (kinase D-interacting substrate of 220 kDa), previously identified as a downstream target of neurotrophin and ephrin receptors, in dendrite development. We report here that knockdown of ARMS/Kidins220 by in utero electroporation impairs dendritic branching in mouse cerebral cortex, and silencing of ARMS/Kidins220 in primary rat hippocampal neurons results in a significant decrease in the length, number, and complexity of the dendritic arbors. Overexpression of cell surface receptor tyrosine kinases, including TrkB and EphB2, in ARMS/Kidins220-deficient neurons can partially rescue the defective dendritic phenotype. More importantly, we show that PI3K (phosphoinositide-3-kinase)- and Akt-mediated signaling pathway is crucial for ARMS/Kidins220-dependent dendrite development. Furthermore, loss of ARMS/Kidins220 significantly reduced the clustering of EphB2 receptor signaling complex in neurons. Our results collectively suggest that ARMS/Kidins220 is a key player in organizing the signaling complex to transduce the extracellular stimuli to cellular responses during dendrite development.

Critical role of the axonal guidance cue EphrinB2 in lung growth, angiogenesis, and repair.

RATIONALE: Lung diseases characterized by alveolar damage currently lack efficient treatments. The mechanisms contributing to normal and impaired alveolar growth and repair are incompletely understood. Axonal guidance cues (AGC) are molecules that guide the outgrowth of axons to their targets. Among these AGCs, members of the Ephrin family also promote angiogenesis, cell migration, and organogenesis outside the nervous system. The role of Ephrins during alveolar growth and repair is unknown. OBJECTIVES: We hypothesized that EphrinB2 promotes alveolar development and repair. METHODS: We used in vitro and in vivo manipulation of EphrinB2 signaling to assess the role of this AGC during normal and impaired lung development. MEASUREMENTS AND MAIN RESULTS: In vivo EphrinB2 knockdown using intranasal siRNA during the postnatal stage of alveolar development in rats arrested alveolar and vascular growth. In a model of O(2)-induced arrested alveolar growth in newborn rats, air space enlargement, loss of lung capillaries, and pulmonary hypertension were associated with decreased lung EphrinB2 and receptor EphB4 expression. In vitro, EphrinB2 preserved alveolar epithelial cell viability in O(2), decreased O(2)-induced alveolar epithelial cell apoptosis, and accelerated alveolar epithelial cell wound healing, maintained lung microvascular endothelial cell viability, and proliferation and vascular network formation. In vivo, treatment with intranasal EphrinB2 decreased alveolar epithelial and endothelial cell apoptosis, preserved alveolar and vascular growth in hyperoxic rats, and attenuated pulmonary hypertension. CONCLUSION: The AGC EphrinB2 may be a new therapeutic target for lung repair and pulmonary hypertension.

Eph and ephrin signaling in the formation of topographic maps.

The axonal connections between the retina and its midbrain target, the superior colliculus (SC), is mapped topographically, such that the spatial relationships of cell bodies in the retina are maintained when terminating in the SC. Topographic map development uses a Cartesian mapping system such that each axis of the retina is mapped independently. Along the nasal-temporal mapping axis, EphAs and ephrin-As, are graded molecular cues required for topographic mapping while the dorsal-ventral axis is mapped in part via EphB and ephrin-Bs. Because both Ephs and ephrins are cell surface molecules they can signal in the forward and reverse directions. Eph/ephrin signaling leads to changes in cytoskeletal dynamics that lead to actin depolymerization and endocytosis guiding axons via attraction and repulsion.

Eph/ephrin signaling in epidermal differentiation and disease.

Eph receptor tyrosine kinases mediate cell-cell communication by interacting with ephrin ligands residing on adjacent cell surfaces. In doing so, these juxtamembrane signaling complexes provide important contextual information about the cellular microenvironment that helps orchestrate tissue morphogenesis and maintain homeostasis. Eph/ephrin signaling has been implicated in various aspects of mammalian skin physiology, with several members of this large family of receptor tyrosine kinases and their ligands present in the epidermis, hair follicles, sebaceous glands, and underlying dermis. This review focuses on the emerging role of Eph receptors and ephrins in epidermal keratinocytes where they can modulate proliferation, migration, differentiation, and death. The activation of Eph receptors by ephrins at sites of cell-cell contact also appears to play a key role in the maturation of intercellular junctional complexes as keratinocytes move out of the basal layer and differentiate in the suprabasal layers of this stratified, squamous epithelium. Furthermore, alterations in the epidermal Eph/ephrin axis have been associated with cutaneous malignancy, wound healing defects and inflammatory skin conditions. These collective observations suggest that the Eph/ephrin cell-cell communication pathway may be amenable to therapeutic intervention for the purpose of restoring epidermal tissue homeostasis and integrity in dermatological disorders.