Why is Fam83h critical for enamel formation?

牙釉质形成的遗传性疾病被归类为 成釉细胞（AI）。约25%的釉质发育异常（AI）病例是由 通过编码四种釉质细胞外基质蛋白的基因，AMELX，ENAM，MMP 20和 KLK 4.最近，我们在六个不同的家庭中发现了导致AI的FAM 83 H突变 患有釉质发育不全症。这表明，FAM 83 H是必要的， 釉质的形成事实上，我们对Fam 83 h蛋白一无所知。我们的目标是 了解Fam 83 h在正常和有缺陷的釉质形成中所起的作用。 假设：由于FAM 83 H突变的人的表型仅限于发展中国家， 牙齿，我们假设Fam 83 H在牙发生过程中表达。因为所有的人工智能- 引起FAM 83 H突变的是显性无义突变，其终止了细胞中的翻译。 最后一个编码外显子（外显子5），我们进一步假设Fam 83 h与其他细胞相互作用， 蛋白质形成功能复合物。我们提出四个具体目标： SA 1：为了表征Fam 83 h表达的时间和空间模式， 并确定其亚细胞定位。 SA 2：分离Fam 83 H蛋白用于结构和功能表征。 SA 3：鉴定Fam 83 H相互作用蛋白。 SA 4：确定截短的Fam 83 h的表达是否干扰釉质形成。 方法：Fam 83 h在牙发生过程中表达的时空模式是 通过原位杂交和免疫组织化学确定。重组和天然 Fam 83 h用于结构和功能研究。Fam 83 h相互作用蛋白的鉴定 使用酵母双杂交系统并使用哺乳动物双杂交系统进行验证， 免疫沉淀和远Western分析。正常和突变的Fam 83 h在大肠杆菌中表达。 成釉细胞作为转基因和它们对釉质形成的影响进行了表征。 健康相关性：患有遗传性釉质缺陷（AI）的患者有疼痛，变形的牙齿， 降低自尊，并认为自己的生活质量较低。发现 FAM 83 H对牙釉质形成至关重要，这是我们理解的一个重大进展 人工智能的病因学这项拟议中的研究可能会导致我们对 成釉细胞内的重要活动和AI新候选基因的发现。

Inherited diseases of dental enamel formation are grouped under the designation of amelogenesis imperfecta (AI). About 25% of all amelogenesis imperfecta (AI) cases are caused by the genes encoding four enamel extracellular matrix proteins, AMELX, ENAM, MMP20, and KLK4. Recently we identified mutations in FAM83H that cause AI in six different families suffering from amelogenesis imperfecta. This demonstrates that FAM83H is necessary for proper enamel formation. Virtually nothing is known about the Fam83h protein. Our objective is to learn the roles played by Fam83h in normal and defective enamel formation. Hypotheses: Because the phenotype in people with FAM83H mutations is limited to developing teeth, we hypothesize that Fam83H is expressed during odontogenesis. Because all of the AI- causing FAM83H mutations are dominant nonsense mutations that terminate translation in the last coding exon (exon 5), we further hypothesize that the Fam83h interacts with other cellular proteins to form functional complexes. We propose four Specific Aims: SA 1: To characterize the temporal and spatial pattern of Fam83h expression during odontogenesis and to determine its subcellular localization. SA 2: To isolate Fam83H protein for structural and functional characterization. SA 3: To identify Fam83H interacting proteins. SA 4: To determine if the expression of truncated Fam83h interferes with amelogenesis. Approach: The temporal and spatial pattern of Fam83h expression during odontogenesis is determined by in situ hybridization and immunohistochemistry. Recombinant and native Fam83h are used in structural and functional studies. Fam83h interacting proteins are identified using the yeast two-hybrid system and validated using a mammalian two-hybrid system, co- immunoprecipitation, and Far Western analyses. Normal and mutated Fam83h are expressed in ameloblasts as transgenes and their effects on enamel formation characterized. Heath Relatedness: Patients with inherited enamel defects (AI) have painful, disfigured teeth, lower self-esteem, and perceive themselves as having an inferior quality of life. The discovery that FAM83H is critical for dental enamel formation is a significant advance in our understanding of the etiology of AI. The proposed research may lead to a breakthrough in our understanding of important activities within ameloblasts and the discovery of new candidate gene(s) for AI.