Mechanism of Xenopus Cranial Neural Crest Cell Migration

描述(由申请人提供)：正确的脑神经脊细胞迁移对于面部、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，但人们对神经脊细胞迁移是如何调控的知之甚少。在非洲爪哇胚胎发育过程中，ADAM 13(一种含有去整合素和金属蛋白酶的蛋白质)的表达与颅神经脊细胞从神经板的外侧边界迁移到腹前站并最终形成面部结构有关(Alfandari等人，1997)。我们正在对表达显性阴性形式的ADAM13的脑神经脊细胞进行的分析表明，ADAM13促进和/或引导它们在三种可能途径中的两种途径迁移。我们的工作假设是，ADAM13切割了一种蛋白质，该蛋白质通常会限制脑神经脊细胞的迁移。这种蛋白可以作为阻止细胞通过的停止信号被插入到迁移路径中，或者被表达在脑神经脊细胞表面，以锚定的形式将细胞固定在适当的位置。为了验证这些假说，并分析其他ADAM和相关的金属蛋白水解酶是否也参与了脑神经脊细胞的迁移，我们提出了以下具体目标。这项提议有三个目的：1)如果缺失ADAM13蛋白的细胞可以使用其他ADAM和相关的金属蛋白酶进行迁移，2)ADAM13是否起到打开迁移途径的“钻头”的作用，3)ADAM13是否切断了将颅神经脊细胞连接到其环境中的锚。使用特定的吗啉寡核苷酸，我们可以阻止包括ADAM13在内的ADAM蛋白在胚胎中的翻译，并测试脑神经脊细胞如何迁移。这可以与ADAM 13功能受阻(使用药物抑制剂)的细胞迁移进行比较。利用移植物，我们将测试缺失ADAM13活性的脑神经脊细胞是否可以跟随具有ADAM13活性的细胞。最后，我们将测试ADAM13是否能够切割已知的将细胞固定下来的蛋白质。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关键一步。此外，有关ADAM对细胞迁移的贡献的信息可能会导致对这些蛋白在各种癌症和转移中的功能的新的理解。特别是，这些蛋白质(ADAM)可能参与了细胞从原始肿瘤逃逸到新位置的过程。

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.