ROLE OF N-GLYCOSYLATION IN E-CADHERIN MEDIATED CELL-CELL ADHESION

N-糖基化在 E-钙粘蛋白介导的细胞粘附中的作用

• 批准号: 7955918
• 负责人: MARIA A. KUKURUZINSKA
• 金额: $ 6.19万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7955918
• 关键词: Adherens Junction; Adhesions; Binding; Biology; Boston; Cadherins; Cell Communication; Cell-Cell Adhesion; Cells; Characteristics; Complex; Computer Retrieval of Information on Scientific Projects Database; Development; Doctor of Philosophy; E-Cadherin; Epithelial Cells; Event; Fingerprint; Funding; Gel; Glycobiology; Grant; Institution; Investigation; Laboratories; Mass Spectrum Analysis; Mediating; Medicine; Membrane Glycoproteins; Membrane Proteins; Methods; Modification; Molecular; Non-Malignant; Pattern; Peptides; Polysaccharides; Protein Binding; Proteins; Proteomics; Research; Research Personnel; Resources; Role; Signal Transduction; Site; Societies; Source; United States National Institutes of Health; Universities; Weight; Western Blotting; Work; cancer cell; cancer type; extracellular; glycosylation; insight; meetings; tumor progression

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. E-cadherin is a 120-kDa membrane glycoprotein expressed in epithelial cells; it is the main player in establishing adherens junctions between cells. The adherens junctions have both intra- and extracellular anchorage points in order to regulate cell-cell interactions.[1-2] Alterations in the assembly or disassembly of adherens junctions occur in association with major changes in the state of cell, including differentiation and proliferation [3], as well as in cancer progression. Loss of E-cadherin function is a frequent event in many types of cancer, commonly caused by diminished or aberrant E-cadherin expression. [4] Results from our own and other laboratories indicate that the N-glycosylation pattern of E-cadherin has a role in the molecular organization of adherens junctions. Specifically, the presence of complex N-glycans is associated with destabilized adherens junctions.[5] Investigation into the structural characteristics of E-cadherin, specifically its N-glycosylation pattern and protein binding partners, provide insight into E-cadherin mediated adhesion and cell signaling functions. Isolation of this large membrane protein has required the development of specific methods. We have now isolated pools of E-cadherin with different binding partners and different N-glycosylation states, depending on cell status. The forms of E-cadherin in malignant cells are more highly glycosylated than are those in non-malignant cells; they migrate at a higher apparent weight, as detected by Western blot and peptide mass fingerprinting of excised SDS-PAGE bands. Proteomic methods have been used to define the composition of the E-cadherin complex under several different conditions. [6] We have found that E-cadherin is associated with destabilizing proteins in malignant cells to a greater extent than when it is present in non-malignant cells. E-cadherin N-glycosylation has also been investigated, specifically site occupancy and N-glycan composition. MALDI-TOF MS and tandem ESI-MS/MS analyses of in-gel enzymatically released N-glycans indicate that E-cadherin contains predominantly complex N-glycans. This work highlights that cell context, the recruitment of specific proteins to the adhesion complex, along with specific structural modifications to E-cadherin may define the overall stability of E-cadherin mediated adhesion. References: 1. Gottardi CJ, Wong E, Gumbiner BM. J Cell Biol 2001; 153(5): 1049-60 2. Perez-Moreno M, Jamora C, Fuchs E. Cell 2003; 112(4): 535-48 3. Gumbiner BM. Nat Rev Mol Cell Biol 2005; 6:622-634 4. Wheelock MJ, Johnson KR. Annu Rev Cell Dev Biol 2003; 19:207-35 5. Liwosz A, Lei T, and Kukuruzinska MA. J Biol Chem 2006; 281:23138-49. 6. Blackmun-Ross K, Kukuruszinska M, Costello CE, 2008, presented at ASMS Conf. and Society for Glycobiology Meeting 7. PhD thesis, K. Blackmun-Ross, Boston University, August 2008.

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 E-钙粘蛋白是上皮细胞中表达的120-kDa膜糖蛋白;它是在细胞之间建立粘附连接的主要参与者。粘附连接具有细胞内和细胞外锚定点以调节细胞-细胞相互作用。[1-2]粘附连接的组装或拆卸的改变与细胞状态的主要变化相关，包括分化和增殖[3]以及癌症进展。在许多类型的癌症中，E-钙粘蛋白功能的丧失是常见的事件，通常由E-钙粘蛋白表达减少或异常引起。[4]从我们自己和其他实验室的结果表明，N-糖基化模式的E-钙粘蛋白有一个作用的分子组织的粘附连接。具体而言，复合N-聚糖的存在与不稳定的粘附连接相关。[5]对E-钙粘蛋白的结构特征，特别是其N-糖基化模式和蛋白结合伴侣的研究，为E-钙粘蛋白介导的粘附和细胞信号传导功能提供了深入了解。 分离这种大的膜蛋白需要开发特定的方法。 我们现在已经分离出了具有不同结合伴侣和不同N-糖基化状态的E-钙粘蛋白库，这取决于细胞状态。 恶性细胞中的E-钙粘蛋白的形式比非恶性细胞中的形式更高度糖基化;它们以更高的表观重量迁移，如通过Western印迹和切下的SDS-PAGE条带的肽质量指纹图谱检测到的。 蛋白质组学方法已用于定义几种不同条件下E-钙粘蛋白复合物的组成。[6]我们已经发现，E-钙粘蛋白与恶性细胞中的不稳定蛋白的相关程度比它存在于非恶性细胞中的程度更大。 还研究了E-钙粘蛋白N-糖基化，特别是位点占用和N-聚糖组成。凝胶内酶促释放N-聚糖的MALDI-TOF MS和串联ESI-MS/MS分析表明，E-钙粘蛋白主要含有复合N-聚糖。 这项工作强调了细胞环境，特定蛋白质的粘附复合物的募集，沿着特定的结构修饰，E-钙粘蛋白可以定义E-钙粘蛋白介导的粘附的整体稳定性。 参考文献： 1. Gottardi CJ，Wong E，Gumbiner BM.细胞生物学杂志2001; 153（5）：1049-60 2.放大图片作者：Perez-Moreno M，Jamora C，Fuchs E. Cell 2003; 112（4）：535-48 3.甘比纳Nat Rev Mol Cell Biol 2005; 6：622-634 4.惠洛克MJ，约翰逊KR. Annu Rev Cell Dev Biol 2003; 19：207-35 5. Liwosz A，Lei T和Kukuruzinska MA. J Biol Chem 2006; 281：23138-49. 6. Blackmun-Ross K，Kukuruszinska M，Costello CE，2008年，在ASMS会议和糖生物学学会会议上发表 7.博士论文，K。Blackmun-Ross，波士顿大学，2008年8月。