Molecular mechanism of the formation of the signaling center in the prospective hindbrain of vertebrate early embryos

脊椎动物早期胚胎前脑信号中心形成的分子机制

• 批准号: 17570170
• 负责人: YAMASU Kyo
• 金额: $ 2.43万
• 依托单位: Saitama University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2005
• 资助国家: 日本
• 起止时间: 2005 至 2007
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-17570170/
• 关键词: developmental biology; zebrafish; brain formation; regulation of gene expression; hindbrain; rhombomere; mutagenesis; transgenesis; ゼブラフィシュ; 遺伝子発現機能; 遺伝子発現制御機構

1. The regulatory mechanism of zebrafish hoxb1b, which is expressed in the caudal neural plate to specify the position of rhombomere 1 in the hindbrain, was examined by the reporter analysis, revealing that hoxb1b expression was specifically driven in the neural plate by two downstream regions, d1 and d4. It was found that these two regions regulate hoxb1b in response to retinoic acid (RA) via two retinoic acid responsive regions (RAREs).2. Pou2 is known to be expressed transiently in the midbrain and hindbrain in late gastrulae, allowing for the development of the midbrain-hindbrain boundary (MHB) and hindbrain. Its regulatory mechanism was examined, showing that pou2 is spatially regulated mainly by the upstream 2.1-kb DNA, that the expression is maintained through an autoregulatory loop mediated by the cooperative function of four Pou2-binding Octamer sequences in the 2.1-kb DNA, and that pou2 is repressed by RA through a RARE in the upstream DNA.3. The internal structure of the late MHB enhancer of fgf8 was examined by introducing different deletions, showing that the MHB enhancer is activated broadly from the midbrain to r5 in the hindbrain by the two Pax2 binding sequences therein, and that a 28-bp region in this enhancer represses expression in the midbrain and posterior hindbrain, giving MHBspecific expression of fgf8. It was also found that this MHB enhancer is broadly conserved during vertebrate evolution.4. Mutant screen was conducted with zebrafish treated with ENU, leading to isolation of a mutant line (aa6k) that shows defects in the midbrain and hindbrain. Besides the brain defect, the mutant embryos have anomalies in touch response and significant jaw defects. The linkage analysis localized the mutation resides in chromosome 3, and revealed several close SSLP markers. Possible involvement of several candidate genes in aa6k mutation is now under investigation.

1. 斑马鱼hoxb1b在尾部神经板中表达，以指定菱形核1在后脑中的位置，通过记者分析检查了斑马鱼hoxb1b的调节机制，揭示了hoxb1b在神经板中的表达是由两个下游区域特异性驱动的、d1 和 d4。研究发现，这两个区域通过两个视黄酸反应区（RARE）调节hoxb1b对视黄酸（RA）的反应。2．已知 Pou2 在原肠胚晚期在中脑和后脑短暂表达，从而促进中脑-后脑边界 (MHB) 和后脑的发育。对其调控机制进行了检查，结果表明pou2主要受上游2.1-kb DNA的空间调控，其表达是通过2.1-kb DNA中四个与Pou2结合的八聚体序列的协同功能介导的自动调节环来维持的，并且pou2通过上游DNA中的RARE被RA抑制。3．通过引入不同的缺失来检查fgf8晚期MHB增强子的内部结构，表明MHB增强子被其中的两个Pax2结合序列从中脑到后脑r5广泛激活，并且该增强子中的28-bp区域被广泛激活。增强子抑制中脑和后脑的表达，从而产生 fgf8 的 MHB 特异性表达。还发现该MHB增强子在脊椎动物进化过程中广泛保守。4．使用经 ENU 处理的斑马鱼进行突变体筛选，分离出显示中脑和后脑缺陷的突变体系 (aa6k)。除了大脑缺陷之外，突变胚胎还存在触觉反应异常和明显的下巴缺陷。连锁分析将突变定位于 3 号染色体，并揭示了几个相近的 SSLP 标记。目前正在研究可能涉及 aa6k 突变的几个候选基因。

项目成果

Initial specification of the epibranchial placode in zebrafish embryos depends on the fibroblast growth factor signal

斑马鱼胚胎鳃上基板的初始规格取决于成纤维细胞生长因子信号

• 发表时间: 2007
• 期刊: Developmental Dynamics 236
• 作者: Nikaido;M.;Doi;K.;Shimizu;T.;Hibi;M.;Kikuchi;Y. and Yamasu;K.
• 通讯作者: K.

ゼブラフィッシュFgf8の2種のisoform(Fgf8とFgf8b)の発現と機能の検討

斑马鱼 Fgf8 两种亚型（Fgf8 和 Fgf8b）的表达和功能检查

• 发表时间: 2006
• 作者: 太田 聡;弥益 恭
• 通讯作者: 弥益 恭

中脳後脳境界におけるfgf8遺伝子の発現制御機構は脊椎動物で保存されている

fgf8基因在中脑-后脑边界的表达控制机制在脊椎动物中是保守的

• 发表时间: 2007
• 作者: 井上詞貴;小林大介;武田洋幸;弥益 恭
• 通讯作者: 弥益 恭

目で見る生物学(三訂版)

用眼睛看到的生物学（第三版）

• 发表时间: 2006
• 作者: 石原 勝敏;団 まりな;浅尾 哲朗;山口 征矢;弥益 恭
• 通讯作者: 弥益 恭

ウィルト 発生生物学

枯萎发育生物学

• 发表时间: 2006
• 作者: Fred H.Wilt;Sarah C.Hake(赤坂;大隅;八杉監訳;弥益分担)
• 通讯作者: 弥益分担)