GENETIC ANALYSIS OF MORPHOGENETIC MECHANISM DURING MOUSE DEVELOPMENT

脊椎动物胚胎的发育过程至少在一定程度上受生长因子等分子的调节。我们关注的是tgf - β超家族成员骨形态发生蛋白（BMPs）在小鼠发育过程中的功能。为了揭示BMP信号的功能，我们利用传统的基因靶向技术培养了缺乏BMP IA型受体（Bmpr或Alk3）和激活素IA型受体（Alk2）的突变小鼠。每个受体的失合性导致严重的胚胎致死性，突变胚胎在胚胎期7.5 （Bmpr）或8.5 （Alk2）死亡。对于Bmpr突变胚，我们发现1)未形成中胚层，2)原肠胚形成前的细胞周期延长。对于Alk2突变胚胎，我们发现1)中胚层形成但未完全分化，2)胚胎外区域（未来的胎盘）的Alk2信号传导对原肠胚形成至关重要，3)Alk2突变细胞不能形成心脏或眼睛。这些结果表明，胚胎发生早期的BMP信号对于细胞生长、原肠胚形成和特定器官（如心脏）的形成非常重要。为了在发育后期对这些基因进行功能分析，我们引入了一种新发明的技术，称为组织特异性基因靶向。利用这项技术，我们以骨特异性的方式（针对成熟的成骨细胞）突变了Bmpr。骨特异性Bmpr缺陷小鼠存活，表明组织特异性基因靶向技术可以避免Bmpr突变的胚胎致死性。突变小鼠比正常窝鼠小，骨质钙化不规则，骨基质沉积较少。这些结果首次证明BMP信号是体内正常骨形成所必需的。除骨外，我们计划在以下组织中建立Alk3的组织特异性突变小鼠；神经组织来源于神经嵴细胞、造血器官、软骨和发育中的肢体。- tgf - β，生长因子，胚胎发生，基因靶向，骨形态发生蛋白，神经发育

Developmental process of vertebrate embryos are regulated, at least in part, by secreting molecules such as growth factors. We are focusing on the function of Bone Morphogenetic Proteins (BMPs) that are the members of TGF-beta superfamily during mouse development. To reveal the function of BMP signaling, we have generated a mutant mouse that is deficient for BMP type IA receptor (Bmpr or Alk3) and activin type IA receptor (Alk2) by conventional gene targeting technologies. Nullizygosity of each receptor caused severe embryonic lethality and mutant embryos die at embryonic day 7.5 (Bmpr) or 8.5 (Alk2). For Bmpr mutant embryos, we found that 1) no mesoderm was formed, and 2) cell cycles prior to gastrulation was prolonged. For the Alk2 mutant embryos, we found that 1) mesoderm was formed but not fully differentiated, 2) Alk2 signaling in the extraembryonic region (future placenta) was critical for gastrulation, and 3) Alk2 mutant cells were not capable to contribute heart or eye. These results suggest that BMP signaling at the early stage of embryogenesis is important for cell growth, gastrulation and formation of particular organs such as heart. For the functional analysis of these genes in later stage of development, we introduced a newly invented technology called tissue- specific gene targeting. Using this technology, we mutated Bmpr in bone-specific manner (specific for mature osteoblasts). The bone- specific Bmpr deficient mice were viable indicating we can avoid embryonic lethality of Bmpr mutation by tissue-specific gene targeting technology. Mutant mice were smaller than normal littermate and show irregular calcification and less deposition of bone matrix in their bones. These results are the first evidences that BMP signaling is required for normal bone formation in vivo. We plan to establish tissue-specific mutant mice for Alk3 in following tissues in addition to the bone; neural tissues derived from neural crest cells, hematopoietic organs, cartilage, and developing limb. - TGF-beta, growth factor, embryogenesis, gene targeting, bone morphogenetic protein, neural development