权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Roles of ShcB/ShcC Docking Proteins in Neural Stem Cells and Neuronal Tumors

ShcB/ShcC 对接蛋白在神经干细胞和神经元肿瘤中的作用

基本信息

批准号：
13680888
负责人：
SAKAI Ryuichi
金额：
$ 2.24万
依托单位：
National Cancer Center Research Institute
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13680888/
关键词：
neuroblastoma docking protein Src family kinases tyrosine kinases ALK metastasis gene amplification RNAi 神経神経芽胞腫ノックアウトマウス

项目摘要

ShoC is a family member of Shc docking proteins, which contain a unique PTB-CH1-SH2 modular organization and conduct as substrates of various receptor tyrosine kinases. Recently, we showed that hyperphosphorylated ShcC detected in some of neuroblastorna cell lines, such as NB-39-nu cells, is associated with constitutively activated anaplastic lymphoma kinase (ALK) caused by the gene amplification. The ALK gene amplification was also detected in about 10% of primary human neuroblastomas. Suppression of ALK expression in NB-39-nu cells by siRNA resulted in decreased phosphorylation level of ShcC, inactivation of MAPKIAkt pathway and cell apoptosis suggesting that ALK tyrosine kinase is dominating survival signal of this neuroblastoma line. To investigate the roles of hyperphosphorylated ShcC in neuroblastoma cell lines, we established NB-39-nu cell lines which overexpress wildtype or mutant ShcC proteins. It was demonstrated that cell-survival and differentiation, cell-motility were markedly impaired in the NB-39-nu cells expressing the 3YF mutant of ShcC which blocks ShcC-Grb2 pathway by the dominant-negative fassion. At the same time, activation level of MAPK and Akt was severely suppressed in these cells. On the other hand, cells overexpressing ShcC as well as the 3YF mutant showed decreased transforming ability, such as anchorage independency and in vivo tumorigenicity that might suggest ShcC-specific negative effects. Loss of persistent phosphorylation of pl3OCas in suspension cell culture was observed in both ShcC overexpressing cells and 3YF cells. These results suggest ShcC might negatively regulate an alternative pathway such as the Src family kinase (SFK)-p130cas pathway in addition to the authentic MAPK and Akt pathways.

ShoC是Shc对接蛋白家族成员，其包含独特的pdb - ch1 - sh2模块化组织，并作为各种受体酪氨酸激酶的底物。最近，我们发现在一些神经母细胞系（如NB-39-nu细胞）中检测到高磷酸化的ShcC与基因扩增引起的组成性激活间变性淋巴瘤激酶（ALK）有关。在约10%的原发性人神经母细胞瘤中也检测到ALK基因扩增。siRNA抑制NB-39-nu细胞中ALK的表达，导致ShcC磷酸化水平降低，MAPKIAkt通路失活，细胞凋亡，提示ALK酪氨酸激酶在该神经母细胞瘤细胞系的存活信号中起主导作用。为了研究过度磷酸化的ShcC在神经母细胞瘤细胞系中的作用，我们建立了过表达野生型或突变型ShcC蛋白的NB-39-nu细胞系。结果表明，在表达ShcC 3YF突变体的NB-39-nu细胞中，细胞存活、分化、细胞运动明显受损，该突变体以显性负表达方式阻断ShcC- grb2通路。同时，MAPK和Akt的激活水平在这些细胞中被严重抑制。另一方面，过表达ShcC和3YF突变体的细胞表现出较低的转化能力，如锚定独立性和体内致瘤性，这可能表明ShcC特异性的负面作用。悬浮细胞培养中，在ShcC过表达细胞和3YF细胞中均观察到pl3OCas持续磷酸化的缺失。这些结果表明，除了真正的MAPK和Akt途径外，ShcC还可能负调控Src家族激酶(SFK)-p130cas途径等替代途径。