Okajima Tetsuya, Xu Aiguo, Irvine Kenneth D
Howard Hughes Medical Institute, Waksman Institute and Department of Molecular Biology and Biochemistry, Rutgers, The State University of New Jersey, Piscataway, NJ 08854, USA.
J Biol Chem. 2003 Oct 24;278(43):42340-5. doi: 10.1074/jbc.M308687200. Epub 2003 Aug 8.
Notch receptors are glycoproteins that mediate a wide range of developmental processes. Notch is modified in its epidermal growth factor-like domains by the addition of fucose to serine or threonine residues. O-Fucosylation is mediated by protein O-fucosyltransferase 1, and down-regulation of this enzyme by RNA interference or mutation of the Ofut1 gene in Drosophila or by mutation of the Pofut1 gene in mouse prevents Notch signaling. To investigate the molecular basis for the requirement for O-linked fucose on Notch, we assayed the ability of tagged, soluble forms of the Notch extracellular domain to bind to its ligands, Delta and Serrate. Down-regulation of OFUT1 by RNA interference in Notch-secreting cells inhibits both Delta-Notch and Serrate-Notch binding, demonstrating a requirement for O-linked fucose for efficient binding of Notch to its ligands. Conversely, overexpression of OFUT1 in cultured cells increases Serrate-Notch binding but inhibits Delta-Notch binding. These effects of OFUT1 are consistent with the consequences of OFUT1 overexpression on Notch signaling in vivo. Intriguingly, they are also opposite to, and are suppressed by, expression of the glycosyltransferase Fringe, which specifically modifies O-linked fucose. Thus, Notch-ligand interactions are dependent upon both the presence and the type of O-fucose glycans.
Notch受体是介导多种发育过程的糖蛋白。Notch在其表皮生长因子样结构域中通过在丝氨酸或苏氨酸残基上添加岩藻糖而被修饰。O-岩藻糖基化由蛋白O-岩藻糖基转移酶1介导，在果蝇中通过RNA干扰或Ofut1基因的突变，或在小鼠中通过Pofut1基因的突变下调该酶，可阻止Notch信号传导。为了研究Notch上O-连接岩藻糖需求的分子基础，我们检测了带有标签的Notch细胞外结构域可溶性形式与配体Delta和Serrate结合的能力。在分泌Notch的细胞中通过RNA干扰下调OFUT1会抑制Delta-Notch和Serrate-Notch的结合，这表明O-连接岩藻糖对于Notch与其配体的有效结合是必需的。相反，在培养细胞中过表达OFUT1会增加Serrate-Notch的结合，但会抑制Delta-Notch的结合。OFUT1的这些作用与OFUT1在体内过表达对Notch信号传导的影响一致。有趣的是，它们也与糖基转移酶Fringe的表达相反，并被其抑制，Fringe可特异性修饰O-连接的岩藻糖。因此，Notch-配体相互作用取决于O-岩藻糖聚糖的存在及类型。
