Cadherin endocytosis in the cranial neural crest

颅神经嵴中的钙粘蛋白内吞作用

• 批准号: 8805561
• 负责人: LISA A TANEYHILL
• 金额: $ 7.6万
• 依托单位: UNIV OF MARYLAND, COLLEGE PARK
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-05-12 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8805561
• 关键词: Abbreviations; Address; Adherens Junction; Affect; Amino Acids; Binding; Biochemistry; Biological; Biological Assay; Biotinylation; Cadherins; Cell membrane; Cell surface; Cell-Cell Adhesion; Cells; Cellular biology; Cephalic; Chick Embryo; Cues; Cytoplasmic Tail; Data; Development; Disease; Dorsal; Down-Regulation; Embryo; Embryology; Embryonic Development; Endocytosis; Endosomes; Epithelial Cells; Event; Excision; Face; Gene Expression; Generations; Genes; Goals; Health; Heart Part; Human; Human body; Imagery; In Vitro; Inherited; Knowledge; Lead; Lysosomes; Malignant Neoplasms; Mediating; Mission; Modeling; Molecular; Molecular Biology; Mutation; N-Cadherin; Nerve; Neural Crest; Neural Crest Cell; Neural tube; Pathway interactions; Pigments; Play; Population; Process; Proteins; Proteolysis; Proteolytic Processing; Public Health; Publishing; Quality of life; Regulation; Research; Research Design; Role; Skin; Somites; Staging; Structure; Syndrome; System; Testing; Work; base; bone; cell behavior; cell motility; cell type; embryo tissue; epithelial to mesenchymal transition; gene repression; human disease; improved; in vitro Assay; in vitro Model; in vivo; in vivo Model; innovation; insight; malformation; migration; molecular marker; mutant; prevent; public health relevance; response; tissue repair; tool

DESCRIPTION (provided by applicant): Abnormalities that occur during neural crest cell (NCC) development are directly responsible for many human congenital and hereditary malformations, diseases and cancers. Premigratory NCCs exist as adherent epithelial cells in the embryonic dorsal neural tube but later become motile through an epithelial-to- mesenchymal transition (EMT) in response to numerous intrinsic and extrinsic cues. Importantly, this event is analogous in scope to abnormal EMTs that underscore human disease and cancer. Although several similarities exist between these normal and aberrant EMTs, the chief amongst these is the cascade of events initiated by loss of cell surface cadherin proteins and the disassembly of adherens junctions to facilitate cell motility. Work from our lab has shown that loss of Cadherin6B (Cad6B) in premigratory cranial NCCs is crucial for NCC EMT. Cadherin reduction in vitro is mediated both transcriptionally and post-translationally through proteolysis and endocytosis. Our prior studies described comparable mechanisms of transcriptional repression and proteolytic processing of Cad6B in vivo during NCC EMT. Our new preliminary data, however, now reveal that Cad6B possesses putative endocytic motifs and co-localizes with markers of endosomes and lysosomes, both in vitro and in vivo. These initial findings suggest that Cad6B is also regulated post-translationally via endocytosis and, to our knowledge, are the first demonstration of cadherin endocytosis during an in vivo EMT. Based on these data, we hypothesize that Cad6B endocytosis plays a critical function during NCC EMT and migration through removal of Cad6B protein from premigratory NCC plasma membranes, thereby promoting the dismantling of adherens junctions and NCC migration. The Specific Aims of this application are to 1) define how mutations in putative endocytic motifs affect Cad6B internalization and 2) assess effects of disruptions in Cad6B endocytosis on NCC EMT and migration. In Aim 1, we will alter specific amino acids within motifs that may regulate Cad6B endocytosis and assess effects on Cad6B internalization, both in vitro and in vivo. In Aim 2, we will evaluate how disruptions in Cad6B endocytosis impact NCC EMT and migration through in vivo and in vitro cell and molecular assays. The proposed research is innovative because it takes a multi-disciplinary approach that combines embryology, biochemistry, and cell and molecular biology to examine post- translational mechanisms of cadherin regulation during a biologically relevant EMT. These studies are significant because the results will enhance our understanding of mechanisms associated with generating migratory cell types during both normal developmental and aberrant EMTs, and could lead to the development of new treatments for human diseases.

描述（由申请人提供）：神经Crest细胞（NCC）发育过程中发生的异常是直接导致许多人类先天性和遗传畸形，疾病和癌症。迁移前NCC作为胚胎背神经管中的粘附上皮细胞存在，但后来通过上皮到间质转变（EMT）响应众多固有和外部线索而变成运动。重要的是，此事件与强调人类疾病和癌症的异常EMT相似。尽管这些正常和异常EMT之间存在几个相似之处，但其中的主要内容是损失细胞表面钙粘蛋白蛋白引发的事件，以及粘附连接处的拆卸以促进细胞运动。我们实验室的工作表明，在移民前NCCS中，Cadherin6b（CAD6B）的损失对于NCC EMT至关重要。通过蛋白水解和内吞作用在转录和翻译后介导了钙粘着蛋白的减少。我们先前的研究描述了NCC EMT期间CAD6B的转录抑制和蛋白水解处理的可比机制。然而，我们的新初步数据现在表明，CAD6B具有假定的内吞基序，并与内体和溶酶体的标记共定位，包括体外和体内。这些最初的发现表明，CAD6B也通过内吞作用在后翻译后受到调节，据我们所知，Cadherin cadherin内吞作用是在体内EMT期间的首次证明。基于这些数据，我们假设CAD6B内吞作用在NCC EMT期间起着关键功能，并且通过从前移民NCC质膜中去除CAD6B蛋白而迁移，从而促进了拆除附着的连接剂和NCC迁移。本应用的具体目的是：1）定义假定的内吞基序中的突变如何影响CAD6B内在化和2）评估CAD6B内吞作用对NCC EMT和迁移的破坏的影响。在AIM 1中，我们将在基序中改变特定的氨基酸，以调节CAD6B内吞作用并评估对CAD6B内部化的影响，无论是体外和体内的。在AIM 2中，我们将评估CAD6B内吞作用中的破坏如何影响NCC EMT和通过体内和体外细胞和分子测定的迁移。拟议的研究具有创新性，因为它采用了一种多学科的方法，该方法结合了胚胎学，生物化学，细胞和分子生物学，以检查生物学相关的EMT期间钙粘蛋白调节的转化后机制。这些研究很重要，因为结果将增强我们对正常发育和异常EMT期间与产生迁移细胞类型相关的机制的理解，并可能导致人类疾病的新治疗方法的发展。