Roles of NGlycans in Carbohydrate Mediated Cell Adhesion

NGlycans 在碳水化合物介导的细胞粘附中的作用

• 批准号: 6573077
• 负责人: Michiko Fukuda
• 金额: $ 26.5万
• 依托单位: SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-03-15 至 2007-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6573077
• 关键词: Golgi apparatus; carbohydrate biosynthesis; carbohydrate structure; cell adhesion; cell migration; enzyme substrate; flow cytometry; gene expression; gene mutation; gene targeting; genetically modified animals; helper T lymphocyte; human tissue; immunocytochemistry; inborn carbohydrate metabolism disorder; keratan sulfate; laboratory mouse; ligands; mannosidase; neutrophil; polysaccharides; selectins; sulfotransferase; tissue /cell culture; transfection

Project III-The Roles of N-glycans in Carbohydrate-Mediated Cell Adhesion (Michiko N. Fukuda, Ph.D.) Alpha-mannosidase IIx(MX) is an enzyme closely related to the Golgi processing alpha-mannosidase II (MII). To investigate the in vivo role of MX, we generated MX-deficient ,mutant mice, and found that MX null mice were born and grew normally but MX null males are sterile due to ineffective spermatogenesis. Our previous studies on HEMPAS disease and evidence derived from the MII gene knock-out mouse suggested the existence of an enzyme potentially compensating for the MII defect. In order to determine if MX and MII compensate for each other's loss in vivo, we produced MII/MX double gene knock-out mice. MII/MX double null mutants are embryonic lethals, indicating that MX and MII together are essential for N-glycan processing. Using these mutant mice, we will define the roles of MX in N-glycan biosynthesis. By generating conditional MII/MX double null mutants, we will determine the contribution of N-glycans to E- and P-selectin in neutrophils and T helper 1 cells. In addition, because we have identified in sulfotransferase responsible for MCD (macular corneal dystrophy), whose product is structurally similar to the L-selectin ligand, we will also determine if this sulfotransferase is involved in the synthesis of L-selectin ligand, we will also determine if this sulfotransferase is involved in the synthesis of L- selectin ligand in vivo in the mouse. Specific aims are to (1) determine the role of MX in N-glycan in vivo in the mouse. Specific aims are to (1) determine the role of MX in N-glycan biosynthesis, (b) to determine the contribution of N-glycans to the ligand activities of E- and P-selectin, and (3) to determine if mouse I-GlcNAc6ST, a sulfotransferase which also functions as keratan sulfate GlcNAc6ST, is involved in sulfation of L-selectin ligand in the mouse.

项目III-N-聚糖在碳水化合物介导的细胞粘附中的作用（Replanko N。Fukuda博士）α-甘露糖苷酶IIx（MX）是与高尔基体加工α-甘露糖苷酶II（MII）密切相关的酶。为了研究MX在体内的作用，我们产生了MX缺陷的突变小鼠，发现MX缺失小鼠出生和生长正常，但MX缺失雄性由于精子发生无效而不育。我们以前对HEMPAS疾病的研究和来自MII基因敲除小鼠的证据表明存在一种可能补偿MII缺陷的酶。为了确定MX和MII是否在体内补偿彼此的损失，我们产生了MII/MX双基因敲除小鼠。MII/MX双无效突变体是胚胎致死的，表明MX和MII一起对于N-聚糖加工是必需的。利用这些突变小鼠，我们将确定MX在N-聚糖生物合成中的作用。通过产生条件性MII/MX双无效突变体，我们将确定N-聚糖对中性粒细胞和T辅助细胞1中E-和P-选择素的贡献。此外，由于我们已经鉴定了导致MCD（黄斑角膜营养不良）的磺基转移酶，其产物在结构上类似于L-选择素配体，我们还将确定该磺基转移酶是否参与L-选择素配体的合成，我们还将确定该磺基转移酶是否参与小鼠体内L-选择素配体的合成。具体目的是（1）确定MX在小鼠体内N-聚糖中的作用。具体目的是（1）确定MX在N-聚糖生物合成中的作用，（B）确定N-聚糖对E-和P-选择素的配体活性的贡献，和（3）确定小鼠I-GlcNAc 6 ST（一种磺基转移酶，也作为硫酸角质素GlcNAc 6 ST起作用）是否参与小鼠中L-选择素配体的硫酸化。