Roles for Glycosylation in Notch Signaling

描述（由申请人提供）：Notch受体是细胞表面 糖蛋白类，它们通过传递信号来控制细胞命运和细胞分裂， 后生动物。组成性或失调的信号传导导致发育 缺陷和癌症。与Notch结合并刺激信号传导的配体 转导是Delta和Serrate/Jagged。我们已经展示了与 果蝇和哺乳动物的边缘蛋白 β 3 N-乙酰葡糖胺基转移酶活性，并直接作用于Notch， GlcNAc与Notch EGF重复序列上的O-岩藻糖结合，从而调节配体诱导的 陷波信号。在果蝇和哺乳动物细胞中， 自主地抑制Notch对锯齿（锯齿状）的反应， 增强了Notch对Delta的反应我们用了一个跨文化记者 在表达Notch CHO细胞中显示躁狂或精神病边缘的测定 抑制对锯齿蛋白1的反应。我们使用了一组CHO糖基化 突变体，以显示通过条纹抑制Notch信号传导并不需要 复合或杂合N-聚糖或唾液酸，但确实需要 连接到Notch EGF重复序列的O-岩藻糖上的边缘，以及随后的作用， beta4GalT- 1。因此O.岩藻糖基转移酶和β 4GalT-1是新的调节剂 Notch信号使用CHO糖基化突变体，共培养Notch信号传导 和可溶性配体结合试验，以及小鼠突变体，我们提出1）为了鉴定 Jagged和Delta诱导的信号传导所需的不同0-岩藻糖聚糖 通过Notch受体1、2、3和4调节Notch信号传导， 哺乳动物边缘（躁狂，精神病和激进）; 2）确定生化 边缘作用干扰Notch信号传导的机制;以及3）为了鉴定 在小鼠中失活基因对Notch信号传导的体内后果 编码Notch信号的新调节剂。

DESCRIPTION (provided by applicant): Notch receptors are cell surface glycoproteins that transduce signals to control cell fate and cell division in metazoans. Constitutive or disregulated signaling leads to developmental defects and cancer. The ligands that bind to Notch and stimulate signal transduction are Delta and Serrate/Jagged. We have shown in collaboration with others that Drosophila and mammalian fringe proteins have a beta3N-acetylglucosaminyltransferase activity and act directly on Notch to add GIcNAc to O-fucose on Notch EGF repeats, thereby modulating ligand-induced Notch signaling. In Drosophila and mammalian cells, fringe acts cell autonomously to inhibit the Notch response to Serrate(Jagged) while potentiating the Notch response to Delta. We have used a co-culture reporter assay to show that manic or lunatic fringe in Notch expressing CHO cells inhibit the response to Jagged1. We have used a panel of CHO glycosylation mutants to show that inhibition of Notch signaling by fringe does not require complex or hybrid N-glycans or sialic acid but does require the action of fringe on O-fucose attached to Notch EGF repeats, and the subsequent action of beta4GalT- 1. Thus O. fucosyltransferase and beta4GalT-1 are novel modulators of Notch signaling. Using CHO glycosylation mutants, co-culture Notch signaling and soluble ligand binding assays, and mouse mutants, we propose 1 )To identify the different 0-fucose glycans required for Jagged- and Delta-induced signaling by Notch receptors 1, 2, 3 and 4 and for the modulation of Notch signaling by mammalian fringes (manic, lunatic and radical); 2) To determine the biochemical mechanism by which fringe action perturbs Notch signaling; and 3) To identify in vivo consequences to Notch signaling in the mouse of inactivating the genes encoding new modulators of Notch signaling.