Mechanism of Xenopus Cranial Neural Crest Cell Migration

描述（由申请人提供）：适当的颅神经嵴（CNC）细胞迁移对于面部，颌骨及其周围神经系统连接的构建至关重要。尽管这很重要，但人们对神经嵴细胞迁移是如何调节的知之甚少。在非洲爪蟾胚胎发育过程中，adam13（一种含有崩解素和金属蛋白酶的蛋白质）的表达与颅神经嵴细胞从神经板外侧边界向腹前站的迁移有关，在那里它们最终形成面部结构（Alfandari等，1997）。我们对表达ADAM13显性阴性形式的颅神经嵴细胞的持续分析表明，ADAM13通过三种可能的途径中的两种促进和/或指导它们的迁移。我们的工作假设是ADAM13切割了一种通常限制颅神经嵴细胞迁移的蛋白质。该蛋白既可以作为停止信号插入到迁移路径中以阻止细胞传代，也可以在颅神经嵴细胞表面表达以作为锚点固定细胞。为了验证这些假设，并分析其他ADAM和相关金属蛋白酶是否也可能参与颅神经嵴细胞迁移，我们提出以下具体目标。该研究有三个目的：1)缺失ADAM13蛋白的细胞是否可以利用其他ADAM13蛋白和相关金属蛋白酶进行迁移；2)ADAM13是否可以作为“钻头”打开迁移途径；3)ADAM13是否切断了连接颅神经嵴细胞与环境的锚点。利用特定的morpholino寡核苷酸，我们可以阻止包括ADAM13在内的ADAM蛋白在胚胎中的翻译，并测试颅神经嵴细胞如何迁移。这可以与adam13功能被阻断的细胞迁移（使用药物抑制剂）进行比较。通过移植物，我们将测试缺乏ADAM13活性的颅神经嵴细胞是否可以跟随具有ADAM13活性的细胞。最后，我们将测试ADAM13是否可以切割已知的锚定细胞的蛋白质。拟议的研究将增加我们对控制面部正常形成的事件的理解，这是诊断和治疗导致异常发育的条件的重要一步。此外，关于ADAM对细胞迁移的贡献的信息可能会导致对这些蛋白质在各种癌症和转移中的功能的新认识。特别是这些蛋白（ADAM）可能参与细胞从原肿瘤向新部位的逃逸。

DESCRIPTION (provided by applicant): Proper cranial neural crest (CNC) cell migration is essential for the construction of the face, jaws and their peripheral nervous system connections. Despite this importance, little is known about how neural crest cell migration is regulated. During Xenopus embryo development the expression of ADAM 13 (a protein containing A Disintegrin And Metalloprotease) correlates with the migration of the cranial neural crest cells from the lateral border of the neural plate to the ventral anterior station where they eventually form facial structures (Alfandari et al., 1997). Our on-going analyses of cranial neural crest cells expressing a dominant negative form of ADAM13 suggest that ADAM13 promotes and/or directs their migration in two of the three possible pathways. Our working hypothesis is that ADAM13 cleaves a protein that normally restricts cranial neural crest cell migration. This protein may either be inserted in the migration path as a stop signal to prevent cell passage or be expressed at the cranial neural crest cell surface to hold the cells in place as an anchor. To test these hypotheses and analyze whether other ADAM and related metalloproteases may also be involved in cranial neural crest cell migration we propose the following specific Aims. This proposal has three Aims to understand 1) if cells missing ADAM13 protein can use other ADAM and related metalloprotease to migrate, 2) if ADAM13 functions as a "drill" to open migration pathways, 3) if ADAM13 cuts an anchor that attaches cranial neural crest cells to their environment. Using specific morpholino oligonucleotides, we can prevent translation of ADAM proteins including ADAM13 in embryos and test how cranial neural crest cells migrate. This can be compared to the migration of cells in which ADAM 13 function is blocked (using drug inhibitor). Using grafts we will test whether cranial neural crest cells missing ADAM13 activity can follow cells that have ADAM13. Finally, we will test if ADAM13 can cleave proteins that are known to anchor cells down. The proposed studies will increase our understanding of events that govern normal formation of the face, an essential step towards diagnosing and treating conditions that lead to abnormal development. Furthermore, information about ADAM contributions to cell migration could lead to new understanding of the function of these proteins in various cancer and metastasis. In particular these protein (ADAM) are likely to be involved in the escape of cells from the original tumor to new sites.