Signaling mechanisms activated by engagement of NGE-PG

NG2蛋白多糖有助于细胞在被VI型胶原覆盖的表面上的扩散和迁移。蛋白多糖的胞质结构域是NG2依赖的扩散和迁移组件所必需的，这表明该结构域参与了影响细胞骨架机械重排的信号转导。细胞铺展在涂有抗NG2单抗的表面，现在被用来研究底物与NG2结合时激活的特定信号机制。这项申请提出的实验旨在确定NG2介导的信号机制，并确定这些过程对细胞行为的影响。具体目标1侧重于确定NG2细胞质结构域的结构元件，这些结构元件是细胞扩散所必需的。这些将通过使用携带关键决定因素的缺失和突变的NG2 cDNA转染的细胞来定义。特异性目的2探讨NG2在激活Rho家族GTP酶(Rho、Rac和CDC42)中的作用，这些GTP酶是控制肌动蛋白细胞骨架重排的分子。特定目标3研究了SH3结构域。这些分子可能提供了NG2和细胞内信号通路之间的联系。特定目的4研究NG2的C末端PDZ结合与含有PDZ结构域的细胞质分子的相互作用。含有PDZ结构域的分子，如MUPPI、FAP-1和Rh亲和素，可能为信号通路提供额外的链接，也可以作为支架将NG2物理定位到关键的细胞微域。

The NG2 proteoglycan contributes to cell spreading and migration on surfaces coated with type VI collagen. The cytoplasmic domain of the proteoglycan is required for the NG2-dependent component of spreading and migration, suggesting that this domain is involved in transducing signals that influence rearrangements of cytoskeletal machinery. Cell spreading on surface coated with anti-NG2 monoclonal antibodies is now being used to study specific signaling mechanisms activated upon engaged of NG2 by the substratum. This application proposes experiments designed to identify signaling mechanisms mediated by NG2 and to define the impact of these processes on cell behavior. Specific aim 1 focuses on identifying structural elements of the NG2 cytoplasmic domain that are required for cell spreading. These will be defined by using cells transfected with NG2 cDNAs carrying deletions and mutations in key determinants. Specific aim 2 probes the role of NG2 in activating rho family GTPases (rho, rac, and cdc42), molecules that control rearrangements of the actin cytoskeleton. Specific aim 3 examines the SH3 domains. These molecules may provide the link between NG2 and intracellular signaling pathways. Specific aim 4 investigates the interaction of the C-terminal PDZ-binding of NG2 with cytoplasmic molecules that contain PDZ domains. PDZ domain-containing molecules such as MUPPI, FAP-1 and rhophilin may provide additional links to signaling pathways, as well as serving as scaffolds for the physical localization of NG2 to critical cellular microdomains.