Molecular Mechanism of Retinoic Acid-Induced Pattern Duplication During Limb Development

视黄酸诱导肢体发育过程中模式复制的分子机制

• 批准号: 04833026
• 负责人: NOHNO Tsutomu
• 金额: $ 1.34万
• 依托单位: KAWASAKI MEDICAL SCHOOL
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1993
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-04833026/
• 关键词: Retinoic acid; Pattern formation; Limb development; Homeobox gene; Growth factor; FGF; TGF-beta; Hepatocyte growth factor; TGF-beta; 成長因子; 膜レセプター; 遺伝子発現

Anterior-posterior pattern of the limb is known to be determined by a signal emanating from the zone of polarizing activity(ZPA) located in the posterior margin of limb bud. Retinoic acid was postulated to be a morphogen of the ZPA.However, recent findings on the mechanism of limb development argue that retinoic acid is an inducer of polarizing activity but not a direct ZPA signal. Mechanism of retinoic acid-induced limb duplication was investigated using chick limb bud, and following results were obtained.1. Expression of HoxD induced by ZPA implantation to the anterior region of the limb bud is distal to the graft and subjacent to the apical ectodermal ridge(AER). Removal of the AER in the posterior region results in the decrease in HoxD expression. Signaling factor from the AER is therefore required for the induction and maintenance of HoxD expression in addition to the ZPA factor. A msh-related homeobox gene, Msx1, is expressed in the AER and underlying distal mesenchyme of the lim … More b bud. Expression of the Msx1 gene depends on the signals from the AER, whereas mesodermal expression of Prx1, a homeobox gene related to Drosophila paired, is independent of the AER.These homeobox genes expressed specifically in the limb bud are differentially regulated by signals from the AER and ZPA.2. Several growth factors, including fibroblast growth factor(FGF) and transforming growth factor beta(TGF-beta), are presumed to be involved in limb development, since the growth factors are known to play important roles for mesoderm induction and axial determination in Xenopus emgbryogenesis.(1) Four FGF receptor genes have been identified, and are expressed in the limb bud. Among them, FGFR2 reveals ectodermal expression and centrally restricted mesodermal expression, suggesting the involvement of the FGF family in skin and bone morphogenesis. The FGF-4 gene is also expressed in the AER of the chick limb bud, as originally observed in the mouse limb bud in organ culture. Ectopic expression of the FGF-4 gene using retrovirus vector results in the supernumeral digits in the chick embryo. Thus, FGF-4 is likely to be one of the AER factors responsible for limb outgrowth.(2) Type I and II receptors for TGF-beta family are known to have characteristic intracellular Ser/Thr kinase domain. Two chicken activin receptors were identified, and the receptor genes are weakly expressed in the early limb bud. Later, the receptor genes are expressed in the prechondrium, myoblasts and ectoderm. Two novel members of the type I receptors were identified in the chick ; one for BMP-2/4 and the other for TGF-beta1. The former is weakly expressed in the prechondrium of the developing limb.(3) Hepatocyte growth factor(HGF) is known to induce secondary body axis in the chick embryo. HGF-producing MRC-5 cells were shown to induce duplicated limb when implanted to the anterior region of the leg bud at budding stages. The HGF gene is expressed in the distal mesenchyme of the limb bud underneath the AER, suggesting HGF as an AER-inducing and/or maintenance factor during limb development. Less

已知肢体的前后模式是由位于肢芽后缘的极化活动区（ZPA）发出的信号确定的。视黄酸被认为是 ZPA 的形态发生素。然而，最近关于肢体发育机制的发现表明，视黄酸是极化活性的诱导剂，但不是直接的 ZPA 信号。利用鸡肢芽研究了视黄酸诱导肢体复制的机制，得到以下结果： 1．由 ZPA 植入肢芽前部区域诱导的 HoxD 表达位于移植物远端且位于顶端外胚层脊 (AER) 下方。去除后部区域的 AER 会导致 HoxD 表达减少。因此，除了 ZPA 因子外，还需要来自 AER 的信号因子来诱导和维持 HoxD 表达。与 msh 相关的同源框基因 Msx1 在 AER 和 lim b 芽的底层远端间充质中表达。 Msx1基因的表达依赖于来自AER的信号，而与果蝇配对相关的同源盒基因Prx1的中胚层表达独立于AER。这些在肢芽中特异表达的同源盒基因受到来自AER和ZPA的信号的差异调节。2。成纤维细胞生长因子（FGF）和转化生长因子β（TGF-β）等多种生长因子被认为参与了肢的发育，因为已知生长因子在非洲爪蟾胚胎发生中中胚层诱导和轴向决定中发挥重要作用。（1）已鉴定出四种FGF受体基因，并在肢芽中表达。其中，FGFR2揭示了外胚层表达和集中限制的中胚层表达，表明FGF家族参与皮肤和骨形态发生。 FGF-4 基因也在鸡肢芽的 AER 中表达，正如最初在器官培养中的小鼠肢芽中观察到的那样。使用逆转录病毒载体异位表达 FGF-4 基因会在鸡胚中产生多余的数字。因此，FGF-4可能是负责肢体生长的AER因子之一。(2)已知TGF-β家族的I型和II型受体具有特征性的细胞内Ser/Thr激酶结构域。鉴定出两种鸡激活素受体，受体基因在早期肢芽中弱表达。随后，受体基因在前软骨细胞、成肌细胞和外胚层中表达。在小鸡身上鉴定出了 I 型受体的两个新成员：一个用于 BMP-2/4，另一个用于 TGF-β1。前者在肢体发育的软骨前膜中微弱表达。(3)已知肝细胞生长因子(HGF)在鸡胚中诱导次级体轴。当在出芽阶段将产生 HGF 的 MRC-5 细胞植入腿芽的前部区域时，可以诱导复制肢体。 HGF 基因在 AER 下方的肢芽远端间质中表达，表明 HGF 在肢体发育过程中是 AER 诱导和/或维持因子。较少的

项目成果

濃野勉,野地澄清: "Serum response factorの補助因子として働くホメオドメインタンパク質" 生化学. 65. 1327-1332 (1993)

Tsutomu Nono，Sumiyo Noji：“同源结构域蛋白作为血清反应因子的辅助因子”生物化学 65. 1327-1332 (1993)。

Koyama, E., Nohno, T., Myokai, F., et al.: "Cooperative activation of Chox-4 homeobox genes by factors from the polarizing region and the apical ridge in chick limb morphogenesis" "Limb Development and Regeneration", Part A, (Fallon, J.F., Goetink, P.F.,

Koyama, E.、Nohno, T.、Myokai, F. 等人：“鸡肢形态发生中来自极化区域和顶脊的因素对 Chox-4 同源盒基因的协同激活”“肢体发育和再生”，

Koyama,E.,Noji,S.,Nohno,T.,et al.: "Cooperative activation of Chox-4 homeobox genes by factors from the polarizing region and the apical ridge during chick limb development" Development,Growth and Differentiation. (1993)

Koyama,E.,Noji,S.,Nohno,T.,et al.：“鸡肢发育过程中极化区和顶脊因子对 Chox-4 同源盒基因的协同激活”发育、生长和分化。

濃野勉,野地澄清: "レチノイン酸による遺伝子発現の調節-その特異的で柔軟な応答のメカニズム" 医学のあゆみ. 163. 891 (1992)

Tsutomu Nono、Sumiyoshi Noji：“视黄酸对基因表达的调节 - 其特异性和灵活的反应机制”《医学史》163. 891 (1992)。

Nohno,T.,Sumitomo,S.,Ishikawa,T.,et al.: "Nucleotide sequence of a cDNA encoding the chicken receptor protein kinase of the TGF-β receptor family" DNA Sequence. 3. 393-396 (1993)

Nohno, T.、Sumitomo, S.、Ishikawa, T. 等：“编码 TGF-β 受体家族的鸡受体蛋白激酶的 cDNA 的核苷酸序列”DNA 序列。