CELL SUBSTRATE INTERACTION IN CRANIOFACIAL MORPHOGENESIS

胚胎形态发生与头颈部发育 是复杂的细胞迁移的结果，形成了 几乎所有的骨骼、肌肉和腺体结构。这个 这些细胞迁移背后的机制还知之甚少， 尽管它们导致了许多常见的先天性 畸形。 最近，我们发现了一种新的间质形成的酶机制。 细胞在基膜基质上的迁移。结果表明，细胞 表面半乳糖基转移酶(GalTase)识别并结合其 基膜促进细胞的适宜底物：底物 黏附和随后的迁移。细胞迁移可被抑制或 用选择性抑制或刺激的试剂刺激， 分别测定表面半乳糖苷酶活性。有趣的是，这些试剂 对细胞在纤维连接蛋白底物上的迁移没有影响，显示底物 GalTase作用的特异性。在本提案中，我们将定义 细胞表面半乳糖酶在迁移过程中的受体功能及其鉴定 基膜中的互补配体。 为了完成这些研究，我们将检查单一专门化的影响 抗半乳糖苷酶抗体对细胞体外迁移的多种影响 底物。实验将定义本地化、周转和 表面半乳糖酶在迁移过程中的循环及其关系 到细胞骨架。抗-GalTase免疫球蛋白将注射到迁移者体内 颅神经脊细胞扰乱颅面形态发生的途径 在活体内。基膜中的GalTase底物将通过 常规的生化技术。迁移将在惰性的基础上进行分析 用纯化的GalTase底物衍生的底物 酶修饰的。因为表面GalTase似乎能识别 层粘连蛋白，我们将研究GalTase和层粘连蛋白之间的关系 受体。作为一个独立的屏幕，CELL：矩阵的交互将是 鉴定为光活化异双功能交联剂。最后， 我们将尝试纠正致命的迁移缺陷T/T突变 体外和体内通过抑制过剩的表面半乳糖苷酶活性降至 正常水平。这些结果将使我们能够定义特定的分子 某些细胞的机制：形态发生和意志过程中的基质相互作用 帮助我们了解先天畸形。

Embryonic morphogenesis, and development of the head and neck in particular, is the result of sophisticated cellular migrations which form virtually all of the skeletal, muscular and glandular structures. The mechanisms that underlie these cell migrations are poorly understood, despite the fact that they are responsible for many common congenital malformations. Recently, we have identified a novel enzymatic mechanism for mesenchymal cell migration on basal lamina substrates. Results suggest that cell surface galactosyltransferase (GalTase) recognizes and binds its appropriate substrates in the basal lamina facilitating cell:substrate adhesion and subsequent migration. Cell migration can be inhibited or stimulated with reagents that selectively inhibit or stimulate, respectively, surface GalTase activity. Interestingly, these reagents have no effect on cell migration on fibronectin substrates, showing substrate specificity of GalTase action. In this proposal, we will define the receptor function of surface GalTase during migration and identify its complementary ligand in the basal lamina. To accomplish these studies, we will examine the effects of monospecific anti-GalTase IgG on cell migration in vitro on a variety of defined substrates. Experiments will define the localization., turnover and recycling of surface GalTase during migration, as well as its relationship to the cytoskeleton. Anti-GalTase IgG will be injected into the migratory pathway of cranial neural crest cells to perturb craniofacial morphogenesis in vivo. The GalTase substrates in the basal lamina will be identified by conventional biochemical techniques. Migration will be analyzed on inert substrates derivatized with purified GalTase substrates that have been enzymatically modified. Since the surface GalTase appears to recognize laminin, we will examine the relationship between GalTase and the laminin receptor. As an independent screen, cell:matrix interactions will be identified with photoactivated heterobifunctional crosslinkers. Finally, we will attempt to correct the lethal migration-deficient T/T mutation in vitro and in vivo by inhibiting the excess surface GalTase activity down to normal levels. These results will enable us to define a specific molecular mechanism for some cell:matrix interactions during morphogenesis and will aid in our understanding of congenital malformations.