Signaling Mechanisms of EphrinB1 in Cell Adhesion, Migration and Invasion

EphrinB1 在细胞粘附、迁移和侵袭中的信号机制

• 批准号: 9343751
• 负责人: Ira Daar
• 金额: $ 68.52万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9343751
• 关键词: Adhesions; Affect; Binding; Biochemical; Biological Models; Breast; Cancer cell line; Cell Adhesion; Cell Fate Control; Cell Line; Cell Lineage; Cell physiology; Cell-Cell Adhesion; Cells; Colon; Data; Defect; Development; Developmental Process; Disease; Disseminated Malignant Neoplasm; Distal; Ectoderm; Embryo; Embryonic Development; Environment; Eph Family Receptors; Ephrins; Epithelial Cells; Event; Family; Family member; Germ Layers; Histologic; Human; Intercellular Junctions; Intervention; Investigation; Knowledge; Laboratories; Ligands; Lung; Maintenance; Malignant Neoplasms; Maps; Mediating; Mesoderm; Metalloproteases; Morphogenesis; Movement; Neoplasm Metastasis; Neural Tube Closure; Neural tube; Neuroblastoma; Ovarian; Pathway interactions; Play; Predisposition; Process; Prostate; Protein Tyrosine Kinase; Proteins; Rana; Regulation; Research; Role; Signal Transduction; Signal Transduction Pathway; Signaling Molecule; System; Tissues; Ubiquitination; Visual Fields; Work; Xenopus; Xenopus oocyte; angiogenesis; cell motility; flotillin; improved; insight; intercellular communication; interest; loss of function; melanoma; member; migration; mutant; neoplastic; planar cell polarity; prevent; receptor; research study; retinal progenitor cell; tumor progression; tumorigenesis; ubiquitin ligase

Our current research interests are aimed toward examining the mechanism by which Eph receptor tyrosine kinases and their ephrin ligands signal events affecting cell-cell adhesion and morphogenetic movements. From the elucidation of these signal transduction pathways we may improve our understanding of oncogenesis. The cell-cell adhesion system plays a major role in normal development and morphogenesis. Inactivation of this adhesion system is thought to play a critical role in cancer invasion and metastasis. The Xenopus embryo is well suited for investigations of these processes because the frog has a well characterized and invariant cell fate map and cell lineage can be easily traced during experiments. Mutant receptors, ligands, and other proteins can be ectopically expressed in embryos. Thus, their effects on signal transduction, motility, and differentiation can be assessed morphologically and histologically as well as biochemically in a developing vertebrate. Our laboratory is currently investigating the role of the Xenopus Eph receptor tyrosine kinases and ephrinB transmembrane ligands in cell signaling and function using the Xenopus oocyte and embryo systems, as well as human cultured cell lines. At present, our emphasis is placed upon the mechanism by which these Eph family members send signals affecting morphogenetic movements. Members of the Eph family have been implicated in regulating numerous developmental processes and have been found to be deregulated in metastatic cancers, for example, prostate, ovarian, breast, colon, neuroblastoma, lung, and melanoma. Our laboratory has continued these studies examining proximal and distal signaling from ephrinB1 that controls cell adhesion and cell movement. We found evidence that ephrinB1 signals via its intracellular domain to control retinal progenitor movement into the eye field by interacting with Dishevelled (dsh), and co-opting the Wnt/planar cell polarity (PCP) pathway. Using biochemical analysis and gain or loss of function experiments, our data suggest that dsh associates with ephrinB1 and mediates ephrinB1 signaling via downstream members of the PCP pathway during eye field formation. Thus, we have used the eye field as a model system for understanding how ephrinB1 controls cell movement. Recently, we have examined the mechanisms by which ephrinB1 affects cell-cell junctions. A body of evidence is emerging that shows a requirement for ephrin ligands in the proper migration of cells, and the formation of cell and tissue boundaries. These processes are dependent on the cell cell adhesion system, which plays a crucial role in normal morphogenetic processes during development, as well as in invasion and metastasis. Although ephrinB ligands are bi- directional signaling molecules, the precise mechanism by which ephrinB1 signals through its intracellular domain to regulate cell-cell adhesion in epithelial cells remains unclear. We demonstrate that the Smurfs regulate tissue separation at mesoderm/ectoderm boundaries through antagonistic interactions with ephrinB1, an Eph receptor ligand that has a key role in regulating the separation of embryonic germ layers. EphrinB1 is targeted by Smurf2 for degradation; however, a Smurf1 interaction with ephrinB1 prevents the association with Smurf2 and precludes ephrinB1 from ubiquitination and degradation, since it is a substantially weaker substrate for Smurf1. Inhibition of Smurf1 expression in embryonic mesoderm results in loss of ephrinB1-mediated separation of this tissue from the ectoderm, which can be rescued by the coincident inhibition of Smurf2 expression. This system of differential interactions between Smurfs and ephrinB1 regulates the maintenance of tissue boundaries through the control of ephrinB protein levels.We believe that these findings significantly broaden our concept and knowledge of the Smurf ubiquitin ligases and ephrinB regulation that affects tissue repulsion and border maintenance during embryogenesis. Furthermore, since both ephrinBs and Smurfs have been strongly implicated in cancer progression, this work may have implications when considering interventions regarding these molecules in metastatic disease. We also have shown, that a decrease in a highly related Eph ligand, ephrinB2 protein, causes neural tube closure defects during Xenopus embryogenesis. Such a decrease in ephrinB2 protein levels is observed on the loss of flotillin-1scaffold protein, a newly identified ephrinB2-binding partner. This dramatic decline in ephrinB2 protein levels on the absence of flotillin-1 expression is specific, and is partly the result of an increased susceptibility to cleavage by the metalloprotease ADAM10. These findings indicate that flotillin-1 regulates ephrinB2 protein levels through ADAM10, and is required for appropriate neural tube morphogenesis in the Xenopus embryo.

我们目前的研究兴趣旨在研究Eph受体酪氨酸激酶及其肝配蛋白配体信号影响细胞-细胞粘附和形态发生运动的机制。通过对这些信号转导通路的阐明，我们可以提高对肿瘤发生的认识。细胞间粘附系统在正常发育和形态发生中发挥着重要作用。这种粘附系统的失活被认为在癌症侵袭和转移中起关键作用。非洲爪蟾胚胎非常适合这些过程的调查，因为青蛙有一个很好的特点和不变的细胞命运图和细胞谱系可以很容易地在实验中跟踪。突变的受体、配体和其他蛋白质可以在胚胎中异位表达。因此，它们对信号转导、运动性和分化的影响可以在发育中的脊椎动物中从形态学、组织学以及生物化学上进行评估。我们的实验室目前正在研究非洲爪蟾Eph受体酪氨酸激酶和ephrinB跨膜配体在细胞信号转导和功能中的作用，使用非洲爪蟾卵母细胞和胚胎系统，以及人类培养的细胞系。目前，我们的重点放在这些Eph家族成员发送影响形态发生运动的信号的机制上。Eph家族的成员已经涉及调节许多发育过程，并且已经发现在转移性癌症例如前列腺癌、卵巢癌、乳腺癌、结肠癌、神经母细胞瘤、肺癌和黑素瘤中失调。我们的实验室继续这些研究检查近端和远端信号ephrinB 1控制细胞粘附和细胞运动。我们发现证据表明ephrinB 1通过其细胞内结构域通过与Dishevelled（dsh）相互作用并选择Wnt/平面细胞极性（PCP）途径来控制视网膜祖细胞运动进入视野。使用生化分析和增益或功能丧失的实验，我们的数据表明，dsh协会与ephrinB 1和介导ephrinB 1信号通过下游成员的PCP途径在视野形成。因此，我们已经使用眼场作为模型系统，了解ephrinB 1如何控制细胞运动。最近，我们研究了ephrinB 1影响细胞-细胞连接的机制。大量的证据表明肝配蛋白配体在细胞的适当迁移以及细胞和组织边界的形成中是必需的。这些过程依赖于细胞粘附系统，该系统在发育过程中的正常形态发生过程以及侵袭和转移中发挥着至关重要的作用。虽然肝配蛋白B配体是双向信号传导分子，但肝配蛋白B1通过其胞内结构域信号传导以调节上皮细胞中的细胞-细胞粘附的精确机制仍不清楚。我们表明，蓝精灵调节组织分离中/外胚层边界通过拮抗相互作用与ephrinB 1，Eph受体配体，具有关键作用，在调节胚胎胚层的分离。EphrinB 1被Smurf 2靶向降解;然而，Smurf 1与EphrinB 1的相互作用阻止了与Smurf 2的结合，并阻止了EphrinB 1的泛素化和降解，因为它是Smurf 1的基本上较弱的底物。Smurf 1在胚胎中胚层表达的抑制导致ephrinB 1介导的该组织从外胚层分离的损失，这可以通过同时抑制Smurf 2表达来挽救。这种系统的差异相互作用之间的蓝精灵和ephrinB 1调节组织边界的维护，通过控制ephrinB的蛋白质levels.We相信，这些研究结果显着拓宽了我们的概念和知识的蓝精灵泛素连接酶和ephrinB调节，影响组织排斥和边界的维持在胚胎发育过程中。此外，由于ephrinBs和Smurfs都与癌症进展密切相关，因此在考虑对转移性疾病中的这些分子进行干预时，这项工作可能会产生影响。我们还表明，在非洲爪蟾胚胎发育过程中，高度相关的Eph配体ephrinB 2蛋白的减少会导致神经管闭合缺陷。这种ephrinB 2蛋白水平的下降是在flotillin-1支架蛋白（一种新鉴定的ephrinB 2结合伴侣）丢失时观察到的。在缺乏flotillin-1表达的情况下，ephrinB 2蛋白水平的这种急剧下降是特异性的，并且部分是对金属蛋白酶ADAM 10切割的敏感性增加的结果。这些发现表明，flotillin-1通过ADAM 10调节ephrinB 2蛋白水平，并需要适当的神经管形态发生在非洲爪蟾胚胎。