ZINC FINGER GENE OP1 IN XENOPUS NEURAL PATTERNING

爪蟾神经模式中的锌指基因 OP1

• 批准号: 6138734
• 负责人: CHRISTA Stefanie MERZDORF
• 金额: $ 1.72万
• 依托单位: MONTANA STATE UNIVERSITY - BOZEMAN
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-08-01 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6138734
• 关键词: Xenopus; chimeric proteins; developmental genetics; gene expression; neurogenesis; protein structure; vertebrate embryology

The goal of this proposal is to understand the molecular basis for vertebrate neural patterning, using the frog Xenopus. Focus will be on the role of the zinc finger gene opl, which was isolated by subtractive cloning in the Sive lab and is one of the earliest known neural markers. Opl is a homologue of the Drosophila pair-rule gene opa which regulates the expression of wingless and engrailed and it is a member of the vertebrate Zic gene family. In mid-gastrula, opl is expressed throughout the prospective neurectoderm with subsequent restriction to the dorsal neural tube and neural crest. Opl can activate expression of the dorsal neural marker pax3, and the midbrain-specific marker engrailed (en-2), suggesting that opl is a regulator of neural patterning. This raises the question of how a single gene can activate genes in different parts of the neural tube. I will address the mechanisms by which this distinction might be made, and the normal role of opl in neural determination. I have shown that opl acts through the Wnt pathway to induce en-2, making opl a strong candidate for an activator of Wnt proteins in vertebrates. In this proposal I will first ask whether induction of pax3 expression by opl also requires Wnt signalling. If it does, candidate Xwnts that may be mediators of pax3 activation will be identified. otx2 is a potential co-regulator of opl and may be involved in activating en-2 expression and preventing pax3 expression in the anterior of the embryo, which will be tested using an animal cap assay. The in vivo activity of opl will be examined using a misexpression assay in whole embryos and by constructing dominant interfering Opl proteins. Together, these approaches will give insight into the function of opl and the mechanism by which it reuglate neural patterning. Since human homologues of opl are highly conserved, these studies will add to our understanding of genes that regulate normal neural tube formation and will suggest mechanisms by which neural tube birth defects may occur.

该提案的目标是利用青蛙爪蟾来了解脊椎动物神经模式的分子基础。 重点将放在锌指基因 opl 的作用上，该基因是在 Sive 实验室通过消减克隆分离出来的，是已知最早的神经标记之一。 Opl是果蝇配对规则基因opa的同源物，调节wingless和engrailed的表达，它是脊椎动物Zic基因家族的成员。 在原肠胚中期，opl 在整个未来的神经外胚层中表达，随后仅限于背侧神经管和神经嵴。 Opl 可以激活背侧神经标记物 pax3 和中脑特异性标记物 engrailed (en-2) 的表达，表明 opl 是神经模式的调节剂。 这就提出了一个问题：单个基因如何激活神经管不同部分的基因。 我将讨论可能产生这种区别的机制，以及 opl 在神经决定中的正常作用。 我已经证明 opl 通过 Wnt 途径诱导 en-2，使 opl 成为脊椎动物中 Wnt 蛋白激活剂的有力候选者。 在这个提案中，我将首先询问 opl 诱导 pax3 表达是否也需要 Wnt 信号传导。 如果确实如此，则可能是 pax3 激活中介的候选 Xwnt 将被识别。 otx2 是 opl 的潜在共同调节因子，可能参与激活 en-2 表达并阻止胚胎前部的 pax3 表达，这将使用动物帽测定进行测试。 将使用整个胚胎中的错误表达测定并通过构建显性干扰 Opl 蛋白来检查 opl 的体内活性。 总之，这些方法将深入了解 opl 的功能及其调节神经模式的机制。 由于 opl 的人类同源物高度保守，这些研究将增进我们对调节正常神经管形成的基因的理解，并提出神经管出生缺陷可能发生的机制。