GLYCAN PROFILES IN HUMAN MYELOID CELLS ASREGULATED BY SIALIDASE ACTIVITY

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 糖链决定簇CD15(也称为Lewis x，或Lex)是人类髓系细胞的识别标志，介导中性粒细胞与树突状细胞的黏附。尽管对这种结构有广泛的兴趣，但CD15表达的机制 仍然是相对没有特征的。因此，萨克斯坦实验室研究了CD15表达增加与人类髓系细胞分化相关的分子基础。对分化细胞的流式细胞仪分析以及使用糖合成和唾液酸酶抑制剂的生化研究表明，CD15表达的增加不是由于CD15的从头生物合成，而是主要由于诱导(2-3)-唾液酸酶活性，该酶从细胞表面唾液酸化CD15(也称为sialyl-Lewis x、Slex或CD15s)产生CD15。这种与分化相关的表面CD15s向CD15的转化主要发生在糖蛋白上。到目前为止，糖基转移酶表达的动态变化是翻译后糖链修饰的唯一原因。这一结果揭示了一种新的范式，它展示了高尔基体膜后糖苷酶活性在关键糖链决定因素的生物合成中的关键作用。在对报告了这些结果的手稿进行评估期间，该资源提供了葡聚糖轮廓，以回应审查者的请求。该资源还为萨克斯坦实验室的博士后克里斯蒂娜·西尔维斯库提供了获得2D-Gel和LC/MS/MS设备的机会，她以前有过质谱学方面的经验。

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. The glycan determinant CD15 (also known as Lewis x, or Lex) is a distinguishing marker for human myeloid cells and mediates neutrophil adhesion to dendritic cells. Despite broad interest in this structure, the mechanisms underlying CD15 expression remain relatively uncharacterized. Accordingly, the Sackstein laboratory has investigated the molecular basis of increasing CD15 expression associated with human myeloid cell differentiation. Flow cytometric analysis of differentiating cells together with biochemical studies using inhibitors of glycan synthesis and of sialidases showed that increased CD15 expression is not due to de novo biosynthesis of CD15, but results predominantly from induction of a(2-3)-sialidase activity, which yields CD15 from cell-surface sialyl-CD15 (also known as sialyl-Lewis x, sLex or CD15s). This differentiation-associated conversion of surface CD15s to CD15 occurs mainly on glycoproteins. Until now, modulation of post-translational glycan modifications has been attributed solely to dynamic variations in glycosyltransferase expression. The results unveiled a new paradigm by demonstrating a critical role for post-Golgi membrane glycosidase activity in the 'biosynthesis' of a key glycan determinant. The Resource provided glycan profiles to respond to a reviewers' request during evaluation of the manuscript which reported these results. The Resource is also providing access to 2D-gel and LC/MS/MS equipment and training for postdoctoral fellow Christina Silvescu from the Sackstein laboratory who has previous experience in mass spectrometry.