STRUCTURE AND BIOSYNTHESIS OF SELECTIN LIGANDS

项目一-选择素配体的结构和生物合成（Minoru Fukuda博士）在之前的资助期内，我们在该领域取得了关键进展。首先，我们发现高内皮小静脉（HEV）中的6-磺基sialyl Lewis x可以在core2分支和延伸的core1 O-聚糖中形成。新型硫转移酶（LSST）和core1延伸酶（core1- beta3glcnact）的表达表明，这些结构在双触角o -聚糖中的表达导致了高活性的l -选择素配体。第二，唾液酸Lewis x在肿瘤细胞上的过度表达导致肿瘤细胞被自然杀伤细胞（NK细胞）溶解，而唾液酸Lewis x的适度表达则有利于肿瘤转移。基于这些发现，我们提出了两个主要的研究方向：1 .阐明HEV周围淋巴结和Peyer's斑块中l -选择素配体的结构和功能。在与博士合作。John Lowe， Michiko Fukuda和Ole Hindsgaul，我们将通过产生突变小鼠来确定HEV外周和粘膜淋巴结中l -选择素配体的完整结构，并确定6-sulfo sialyl Lewis x是l -选择素配体的作用。2。碳水化合物在肿瘤转移中的作用。在与博士合作。Michiko Fukuda和John Lowe，我们将确定sialyl Lewis x的量是否在促进肿瘤转移或NK细胞介导的细胞溶解中起主要作用。我们还将确定从肿瘤细胞中分离的细胞表面碳水化合物是否抑制肿瘤转移或NK细胞介导的细胞溶解。这些研究将使我们了解选择素配体的结构和生物合成，以及肿瘤细胞表面碳水化合物在肿瘤转移和免疫细胞识别中的作用。

Project I- Structure and Biosynthesis of Selectin Ligands (Minoru Fukuda, Ph.D.) In the previous funding period, we have made critical progress in this field. First, we discovered that 6-sulfo sialyl Lewis x in high endothelial venules (HEV) can be formed in core2 branch and extended core1 O- glycans. Expression of novel sulfotransferase (LSST) and core1 extension enzyme (core1-beta3GlcNAcT) demonstrated that the expression of these structures in biantennary O-glycans leads to highly active L-selectin ligands. Second, that excessive expression of sialyl Lewis x on tumor cells results in cytolysis of those tumor cells by nature killer (NK) cells, while moderate expression of sialyl Lewis x facilitates tumor metastasis. Based on these findings, two major areas of study are proposed: I. Elucidation of structure and function of L-selectin ligands in HEV at peripheral lymph nodes and Peyer's patches. In collaboration with Drs. John Lowe, Michiko Fukuda, and Ole Hindsgaul, we will determine the complete structure of L-selectin ligands at HEV in peripheral and mucosal lymph nodes, and determine the roles of 6-sulfo sialyl Lewis x is an L-selectin ligand by generating a mutant mouse. II. Roles of carbohydrates in tumor metastasis. In collaboration with Drs. Michiko Fukuda and John Lowe, we will determine if the amount of sialyl Lewis x plays a major role in facilitating tumor metastasis or NK cell-mediated cytolysis. We will also determine if cell surface carbohydrates isolated from tumor cells inhibit tumor metastasis or NK cell-mediated cytolysis. These studies will allow us to understand the structure and biosynthesis of selectin ligands and the roles of cell surface carbohydrates of tumor cells in tumor metastasis and immune cell recognition.