THE A(1,3)FUCOSYLTRANSFERASE GENES AND SELECTIN LIGAND EXPRESSION

选择素依赖性粘附事件在免疫介导的病原体处置中对白细胞运输提供了重要贡献。选择素依赖性粘附还可有助于将自然杀伤（NK）细胞和其他免疫细胞递送至肿瘤，并可由此促进体内癌细胞的NK细胞依赖性消除。选择素-选择素反受体偶联还可以控制骨髓中的细胞生长和分化，因为选择素配体缺乏与异常的骨髓细胞增殖相关。α（1，3）岩藻糖基转移酶Fuc-TIV和Fuc-TVII催化活性选择配体的制备所必需的岩藻糖基化事件。因此，这些酶能够控制白细胞运输事件和骨髓增殖过程。为了更全面地了解这两种酶如何促进（i）选择素反受体合成，活性和结构，（ii）骨髓细胞增殖和（iii）白细胞运输的分子，提出了四个具体目标。具体目标#1将定义在高内皮微静脉和白细胞中由Fuc-TIV和Fuc-TVII催化合成的聚糖的结构。这项工作将使用放射性标记的单糖，以及选择素配体岩藻糖基化存在组成性或条件性缺陷以及选择素配体活性存在相应缺陷的突变小鼠品系（与Minora Fukuda博士合作，项目I）。具体目标II将定义在髓样分化期间Fuc-TVII以及E-和P-选择素配体活性的表达模式，以开始理解在选择素配体活性不存在的情况下在髓样谱系中观察到的异常增殖。将使用其中已经安装了绿色荧光蛋白基因的小鼠来鉴定表达Fuc TVII的、骨髓来源的髓样祖细胞，并在Fuc-TVII基因座的控制下表达。具体目标#III将探索Fuc TVII和β（1，4）GalNAc转移酶之间的合成相互作用，其可以改变T淋巴细胞谱系中的选择素配体结构和活性。这些研究将使用缺失β（1，4）GalNAc基因座的小鼠和使用蓝色荧光蛋白基因已安装在其中并表达控制β（1，4）GalNAc基因座的小鼠完成。聚糖结构分析将补充这些研究（与Minoru Fukuda博士合作，项目I）。具体目标#IV将确定选择素配体和其他粘附分子在NK细胞体外和体内杀死癌细胞、体内运输至肿瘤和体内抑制肿瘤生长的能力中的作用。

Selectin-dependent adhesive events provide essential contributions to leukocyte trafficking in immune-mediated disposition of pathogens. Selectin-dependent adhesion may also contribute to delivery of natural killer (NK) cells and other immune cells to tumors, and may thereby facilitate NK cell-dependent elimination of cancer cells in vivo. The selectin-selectin counter-receptor couple may also control cell growth and differentiation in the marrow, since selectin ligand deficiencies are associated with aberrant myeloid cell proliferation. The alpha (1,3) fucosyltransferases Fuc-TIV and Fuc-TVII catalyze fucosylation events essential to the elaboration of active selecting ligands. These enzymes are thus in a position to control leukocyte trafficking events, and myeloid proliferation processes. To more completely understand the molecular of how these two enzymes contribute to (i) selectin counter-receptor synthesis, activity, and structure, (ii) myeloid cell proliferation, and (iii) leukocyte trafficking, four specific aims are proposed. Specific Aim #1 will define the structures of the glycans whose synthesis is catalyzed by Fuc-TIV and Fuc-TVII, in high endothelial venules, and in leukocytes. This work will use radiolabeled monosaccharides, and strains of mutant mice with constitutive or conditional defects in fucosylation of selectin ligands and corresponding defects in selectin ligand activity (collaboration with Dr. Minora Fukuda, Project I). Specific Aim #II will define the expression patterns of Fuc-TVII, and E- and P-selectin ligand activities, during myeloid differentiation, to begin to understand the aberrant proliferation observed in the myeloid lineage in the absence of selectin ligand activity Fuc TVII-expressing, marrow-derived myeloid progenitors will be identified using mice where the green fluorescence protein gene has been installed within, and is expressed under the control of the Fuc-TVII locus. Specific Aim #III will explore a synthetic interaction between Fuc TVII and a beta (1,4) GalNAc transferase that may modify selectin ligand structure and activity in the T lymphocyte lineage. These studies will be completed using mice deleted for the beta (1,4)GalNAc locus, and using mice where the blue fluorescence protein gene has been installed within, and is expressed control the control of the beta (1,4)GalNAc locus. Glycan structural analyses will complement these studies (collaboration with Dr. Minoru Fukuda, Project I). Specific Aim #IV will determine the role of selectin ligands, and other adhesion molecules, in the ability of NK cells to kill cancer cell in vitro and in vivo, to traffic to tumors in vivo, and to suppress tumor growth in vivo.