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Functional Regulation of Erb B family by N-glycosylation

N-糖基化对 Erb B 家族的功能调节

基本信息

批准号：
18590272
负责人：
TAKAHASHI Motoko
金额：
$ 2.11万
依托单位：
Sapporo Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18590272/
关键词：
N-glycan glycosylation ErbB EGFR dimerization adenylyl cyclase 糖鎖増殖因子受容体 ErbB2 ErbB3 二量体形成

项目摘要

Glycosylation is one of the most common post-translational modification, and is known to change physico-chemical properties of proteins. In this study, I examine the function of N-glycan of ErbB family and adenylyl cyclase.ErbB3 has 10 potential glycosylation sites in its extracellular domain. A series of human ErbB3 molecules devoid of N-glycan were prepared and transfected to Flp-In-CHO cells for stable expression. It was revealed that the Asn 418 to Gln mutant (N418Q) mutant of ErbB3 underwent ligand-independent homodimerization. When overexpressed with ErbB2, ligand-independent hetero-dimerization was observed and Erk and Akt phosphorylation was upregulated in the absence of of ErbB3 coexpressed with ErbB2 promoted cell proliferation and colony formation in soft agar in an Erk and Akt-dependent manner. The N418Q mutation also promoted the growth of tumors in athymic mice when injected subcutaneously. Thus, Asn 418-linked N-glycan in ErbB3 plays an essential role in regulating receptor heterodimerization with ErbB2 and might have an effect on transforming activity.Adenylyl cyclase III (ACIII) has 2 potential glycosylation sites in its extracellular domain, and it was suggested that glycosylation is involved in its enzymatic activity. When processing of glycosylation was suppressed by introducing GnT-III, one of glycosyltransferases, forskolin-induced ACIII activation and downstream signaling such as the synthesis of cAMP and the phosphorylation of CREB were significantly upregulated. Now the mechanisms of the regulation of ACIII by the structure of N-glycan is being investigated.

糖基化是最常见的翻译后修饰之一，并且已知改变蛋白质的物理化学性质。本研究对ErbB家族的N-聚糖和腺苷酸环化酶的功能进行了研究。制备了一系列缺乏N-聚糖的人ErbB 3分子，并将其转染至Flp-In-CHO细胞中进行稳定表达。结果表明，ErbB 3的Asn 418到Gln突变体（N418 Q）经历配体非依赖性同源二聚化。当与ErbB 2过表达时，观察到配体非依赖性异源二聚化，并且Erk和Akt磷酸化在不存在ErbB 3的情况下上调，ErbB 3与ErbB 2共表达以Erk和Akt依赖性方式促进细胞增殖和软琼脂中的集落形成。当皮下注射时，N418 Q突变也促进了无胸腺小鼠中肿瘤的生长。腺苷酸环化酶III（Adenylyl cyclase III，ACIII）的胞外域存在2个糖基化位点，提示糖基化参与了其酶活性的调节。当通过引入GnT-III抑制糖基化的加工时，糖基转移酶之一，毛喉素诱导的ACIII活化和下游信号传导如cAMP的合成和CREB的磷酸化显著上调。目前正在研究N-聚糖结构对ACIII的调控机制。