Ethanol-induced Lower Jaw Loss in Bmp Signaling Pathway Mutants

描述(由申请人提供)：胎儿酒精谱系障碍(FASD)描述了一系列由酒精引起的发育缺陷，包括颅面畸形，如下颌骨发育不全。每年大约每100名在美国出生的儿童中就有1人受到影响。虽然胎儿暴露在酒精中会导致FASD，但这些乙醇引起的头面部缺陷背后的遗传学几乎一无所知。颅面的大部分骨骼，包括下颌骨，都是由颅神经脊细胞产生的。神经脊细胞和上皮组织之间的复杂相互作用对颅面形态发生具有重要意义。这些相互作用是由骨形态发生蛋白(BMP)信号通路等协调的。使用斑马鱼的初步数据表明，BMP信号通路的成员是乙醇敏感基因。Bmp2b纯合子突变胚胎在颅面发育开始前死亡，而bmp2b杂合胚胎和Bmp4纯合子突变胚胎(统称为BMP途径突变)经历正常的头面部发育。然而，BMP途径突变体表现出乙醇诱导的下颌骨Meckel‘s软骨的缺陷。鞘氨醇受体S1PR2的突变会导致与Meckel‘s软骨非常相似的缺陷，这是由于前内胚层的形态发生中断以及随后的口腔外胚层缺陷造成的。这一发现表明，假设乙醇扰乱了BMP途径突变体的前内胚层形态发生，导致口腔外胚层缺陷和随后的颅面发育。在本建议的具体目标1中，1)乙醇如何干扰BMP途径突变体在内胚层形态发生过程中的细胞行为将被可视化；2)内胚层和口腔外胚层之间的相互作用将通过产生标记内胚层和口腔外胚层的双重转基因株来直接可视化。在具体目标2中，将通过移植实验和原位标记分析来确定BMP通路信号在下颌骨构型中的作用机制。在具体目标3中，将绘制从正向遗传筛查中恢复的乙醇敏感突变体的图谱，并表征总体的乙醇敏感性。总体而言，这一应用将提供对导致FASD头面部缺陷变异性的基因-乙醇相互作用的更多洞察力。此外，由于斑马鱼和人类之间的基因功能保守，这项工作将直接转化为对人类种群中候选基因的研究，并有助于更好地诊断和治疗FASD。

DESCRIPTION (provided by applicant): Fetal Alcohol Spectrum Disorder (FASD) describes a wide array of ethanol-induced developmental defects, including craniofacial dysmorphology, such as lower jaw hypoplasia. It affects approximately 1 in 100 children born in the United States each year. Although fetal exposure to alcohol causes FASD, virtually nothing of the genetics behind these ethanol-induced craniofacial defects is understood. The majority of the craniofacial skeleton, including the lower jaw, is generated by cranial neural crest cells. Complex interactions between neural crest cells and epithelial tissues are important for craniofacial morphogenesis. These interactions are orchestrated by, among others, the Bone Morphogenic Protein (Bmp) signaling pathway. Preliminary data using zebrafish demonstrate that members of the Bmp signaling pathway are ethanol-sensitive loci. While bmp2b homozygous mutant embryos die before craniofacial development initiates, bmp2b heterozygous embryos and bmp4 homozygous mutant embryos (together termed Bmp-pathway mutants) undergo normal craniofacial development. However, Bmp-pathway mutants display ethanol-induced defects to Meckel's cartilage the lower jaw. Mutation of the sphingosine receptor, s1pr2, causes strikingly similar defects to Meckel's cartilage due to disrupted morphogenesis of the anterior endoderm and subsequent defects to the oral ectoderm. The findings suggest the hypothesis that ethanol disrupts anterior endoderm morphogenesis in Bmp-pathway mutants resulting in defects in the oral ectoderm and subsequent craniofacial development. In Specific Aim 1 of this proposal, 1) how ethanol perturbs cell behaviors during endodermal morphogenesis in Bmp-pathway mutants will be visualized and 2) the interaction between the endoderm and oral ectoderm will be directly visualized through generation of a double transgenic line that labels the endoderm and the oral ectoderm. In Specific Aim 2, the mechanism of Bmp-pathway signaling in patterning the lower jaw will be determined through transplantation experiments and in situ marker analyses. In Specific Aim 3, an ethanol- sensitive mutant recovered from a forward genetic screen will be mapped and the overall ethanol sensitivity characterized. Overall, this application will provide greater insight into the gene-ethanol interactions that lead to the variability of craniofacial defects in FASD. In addition, because of the conservation of gene function between zebrafish and humans, this work will directly translate to studies of candidate genes in human populations and allow better for diagnosis and treatment of FASD.