LAMININ-5 AND HEMIDESMOSOMES IN ORAL EPITHELIAL CELLS

口腔上皮细胞中的层粘连蛋白-5 和半桥粒

• 批准号: 6323346
• 负责人: JONATHAN C. JONES
• 金额: $ 19.56万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-06-01 至 2001-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6323346
• 关键词: cell adhesion; cytoskeleton; epithelium; extracellular matrix; gingiva; integrins; intercellular connection; intermolecular interaction; laminin; molecular cloning; oral mucosa; protein isoforms; recombinant proteins; tissue /cell culture

Hemidesmosomes, cell-matrix connector devices, are used by oral epithelial to attach to the connective tissue of the gingiva as well as the tooth surface. In the cytoplasm of an oral epithelial cell hemidesmosomes are also sites of cytoskeleton anchorage to the cell surface. During the course of periodontal disease, there is, of necessity, modulation in hemidesmosome integrity as epithelial cells migrate along the tooth to form the "long junctional epithelium". Before we can study the role of hemidesmosomes in the pathogenesis of periodontal disease we must have a basic understanding of the molecular components involved in both the assembly and structural maintenance of this complex morphological entity. To this end, in Project 1 of this program project application, we plan to focus our efforts on functional analyses of an adhesive extracellular glycoprotein component of hemidesmosomes called laminin-5 and its impact on oral epithelial cells. In Aim I we address the identification of domains of the laminin-5 heterotrimer involved in nucleation of hemidesmosome assembly. This will involve preparation of recombinant laminin-5 proteins as well as laminin-5 peptides. In Aim 2, these recombinant proteins and peptides will be assayed for their ability a) to induce rapid adhesion of oral epithelial cells and b) to nucleate assembly of hemidesmosomes. As alternative strategies for identification of functional important domains of laminin-5, in Aim 2 we will use peptides to compete the adhesive properties of intact laminin-5. In Aim 3, we wish to identify the substrate binding site(s) of laminin-5. To this end, proteolytic fragments of laminin-5 will be assayed for a) their ability to adhere to surfaces and/or b) to compete with native laminin-5 for substrate binding. Functionally active fragments will be further characterized by microsequencing. In Aim 4, the impact of cell binding to laminin- 5 on hemidesmosome components will be assayed biochemically. We are particularly interested in the possibility that laminin-5 plays a role in matrix-cell signaling via phosphorylation events. Finally, in Aim 5, we wish to identify membrane interactions of laminin-5. This linkage is necessary for the stabilization of the oral epithelium on the tooth surface.

Hemidesmosmosmes，Cell-Matrix连接器设备，由口服使用 上皮也附着在牙龈的结缔组织上 作为牙齿表面。 在口腔上皮细胞的细胞质中 Hemidesmosomes也是细胞骨架锚定部位的部位 细胞表面。在牙周疾病过程中，有 必要性，作为上皮的下半体小体完整性的调制 细胞沿牙齿迁移，形成“长连接 上皮”。在我们可以研究半体体在 牙周疾病的发病机理我们必须有基本的 了解涉及的分子成分 该综合体的组装和结构维护 形态实体。 为此，在该计划的项目1中 项目应用程序，我们计划将精力集中在功能上 分析细胞外糖蛋白成分的分析 Hemidesmosomes称为层粘连蛋白5及其对口服上皮的影响 细胞。 目的我是解决域名的识别 层粘连蛋白-5杂三聚体参与半底体的成核 集会。 这将涉及准备重组层粘连蛋白5 蛋白质以及层粘连蛋白-5肽。 在AIM 2中，这些 重组蛋白和肽的能力将被分析 a）诱导口腔上皮细胞的快速粘附，b） 核心小体的成核组装。 作为替代策略 用于鉴定层粘连蛋白5的功能重要结构域 目的2我们将使用肽来竞争 完整的层粘连蛋白5。 在AIM 3中，我们希望确定基板 层粘连蛋白5的结合位点。 为此，蛋白水解碎片 laminin-5将被分析a）它们粘附在表面的能力 和/或b）与天然层粘连蛋白5竞争底物结合。 功能上活性的片段将进一步表征 微链试验。 在AIM 4中，细胞与层粘连蛋白结合的影响 5上的半体分量将通过生化测定。 我们对laminin-5的可能性特别感兴趣 通过磷酸化事件在基质细胞信号传导中起作用。 最后，在AIM 5中，我们希望确定 层粘连蛋白5。这种联系对于稳定 牙齿表面的口服上皮。