Regulation of Granulopoiesis by Fucosylation-Dependent Notch Signaling

描述(申请人提供)：这项资助申请的总体目标是支持K08职业发展奖，该奖项重点研究岩藻糖化依赖的Notch信号通路在调节髓系命运规范和髓系发展中的作用。岩藻糖化的葡聚糖与许多生物过程有关。在各种类型的细胞命运规范中，作为Notch胞外EGF重复序列上O-连接岩藻糖基的岩藻糖基化是Notch信号转导所必需的。这一建议的首席调查者周兰在她之前与John Lowe博士的合作以及她在凯斯西储大学进行的研究中，在FX-/-小鼠中发现了一种骨髓增殖表型，这种表型在细胞岩藻糖基化方面存在条件缺陷，是由于髓系祖细胞上失去了依赖Notch的信号转导。在这项建议中，周博士将使用转基因Notch报告鼠来检查骨髓造血干细胞和髓系祖细胞是否由于岩藻糖基化缺陷而抑制了Notch信号转导。然后，她将确定岩藻糖基化缺陷的造血祖细胞是否表现出与骨髓基质环境相互作用的改变。她将利用pofut！表达缺陷的小鼠，进一步研究O-岩藻糖化在粒细胞生成中修饰Notch信号的作用。这介导了Notch EGF重复序列的岩藻糖基化，并通过研究Pofutl-/-ES细胞在体外的造血作用。最后，她将具体检查Notch上的哪种O-岩藻糖残留物！EGF重复序列对Notch发挥其粒系调控作用具有重要意义。这将是第一次研究在骨髓微环境中加入岩藻糖化糖蛋白(如Notch)后，骨髓干细胞和祖细胞中激活的控制骨髓生成的信号转导通路，以及最终有助于调节粒细胞生成的机制。这些研究的结果将提供关于O-岩藻糖基化缺陷导致的Notch信号异常是否在人类慢性骨髓增生性疾病中发挥作用的见解。该提案的结果将为未来的研究提供基础，以探索包括Notch和Notch配体在内的岩藻糖化糖蛋白，以及其他分泌或膜相关岩藻糖化多糖在造血干细胞自我更新、髓系命运的指定和确定以及支持造血干细胞活性的骨髓生态位能力中的意义。

DESCRIPTION (provided by applicant): The overall goal of this grant application is to support a K08 career development award that focuses on studying the role of the fucosylation-dependent Notch signaling pathway in the regulation of myeloid fate specification and myeloid lineage development. Fucosylated glycans are implicated in many bilogical processes. Fucosylation as O-linked fucose moieties on Notch extracellular EGF repeats is required for Notch signaling in various types of cell fate specification. Lan Zhou, the principle investigator of this proposal, in her previous work with Dr. John Lowe, and in her research conducted at Case Western Reserve University, identified a myeloproliferative phenotype in FX-/- mice, which are conditionally deficient in cellular fucosylation, is consequent to loss of Notch-dependent signal transduction on myeloid progenitor cells. In this proposal, Dr. Zhou will use a transgenic Notch reporter mouse to examine whether the marrrow hematopoietic stem and myeloid progenitor cells have suppressed Notch signal transduction as a consequence of fucosylation deficiency. She will then determine if fucosylation-deficient hematopoietic progenitor cells display altered interactions with the marrow stromal environment. She will further examine the role of O-fucosylation in modifying Notch signaling in granulopoiesis by using mice deficient in the expression of pofut! that mediates the fucosylation of Notch EGF repeats, and by studying the pofutl-/- ES cell in vitro hematopoiesis. Finally, she will examine specifically which O-fucose residue on Notch! EGF repeats is important for Notch to exert its effect in granulopoietic regulation. This will be the first study to define the signaling transduction pathways controlling myelopoiesis that are activated in bone marrow stem and progenitor cells after engagement of fucosylated glycoprotein, such as Notch, with bone marrow microenviroment, and the mechanisms that ultimately contribute to the regulation of granulopoiesis. The outcome of these studies will provide insights as to whether dysregulated Notch signaling consequent to deficiency of O-fucosylation could play a role in human chronic myeloproliferative diseases. Resutls from the proposal will provide the basis for future studies to explore the significance of fucosylated glycoproteins including Notch and Notch ligands, and, perhaps other secreted or membrane associated fucosylated glycans, in hematopoietic stem cell self-renewal, myeloid fate specification and determination, and bone marrow niche competency in supproting hematopoietic stem cell activity.