Why is Fam83h critical for enamel formation?

为什么 Fam83h 对于牙釉质形成至关重要？

• 批准号: 7623768
• 负责人: JAN Ching Chun HU
• 金额: $ 33.02万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7623768
• 关键词: 8q24.3; Ameloblasts; Amelogenesis; Amelogenesis Imperfecta; Bacteria; Biochemical; Biological; Candidate Disease Gene; Cells; Characteristics; Chromosomes; Chromosomes, Human, Pair 8; Co-Immunoprecipitations; Code; Complex; Defect; Dental Enamel; Development; Disease; Dominant-Negative Mutation; Electron Microscopy; Enamel Formation; Etiology; Exons; Extracellular Matrix Proteins; Family; Genes; Genetic; Goals; Histologic; Immunohistochemistry; In Situ Hybridization; In Vitro; Inferior; Inherited; Lead; Learning; Link; MMP-20; Mass Spectrum Analysis; Monoclonal Antibodies; Mutate; Mutation; Nonsense Mutation; Odontogenesis; Oral health; Pain; Patients; Pattern; Peptides; Phenotype; Phosphoserine; Phosphothreonine; Phosphotyrosine; Play; Post-Translational Protein Processing; Principal Investigator; Proteins; Quality of life; Recombinants; Research; Role; Scanning; Secondary to; Structural Protein; Structure; System; Testing; Time Study; Tissues; Tooth structure; Transgenes; Transgenic Mice; Translations; Western Blotting; amelogenin; base; enamelin; genome-wide; glycosylation; in vivo; light microscopy; malformation; member; mouse model; novel; polyclonal antibody; programs; public health relevance; self esteem; yeast two hybrid system

DESCRIPTION (provided by applicant): 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. PUBLIC HEALTH RELEVANCE: Our group has determined that defects in FAM83H (family with sequence similarity 83 member H) on chromosome 8 cause severe enamel malformations (autosomal dominant hypocalcification amelogenesis imperfecta). Our objectives are to discover the normal function of Fam83h and how defects in this protein result in enamel malformations. Our long term goal is to advance understanding of normal and pathological tooth formation to stimulate improvements in the public's oral health.

描述（由申请人提供）：牙齿牙釉质形成的遗传疾病被分组为未植物的指定Imperfecta（AI）。在所有未蛋白质发生Imperfecta（AI）病例中，约有25％是由编码四个搪瓷细胞外基质蛋白的基因引起的，Amelx，Enam，MMP20和KLK4。最近，我们确定了FAM83H中引起AI的突变，该突变患有六个不同的家庭，患有无肿瘤发生的不完美。这表明FAM83H对于适当的搪瓷形成是必要的。 FAM83H蛋白几乎一无所知。我们的目标是学习FAM83H在正常和有缺陷的搪瓷形成中扮演的角色。假设：由于患有FAM83H突变的人的表型仅限于发育牙齿，因此我们假设FAM83H在牙肠发生过程中表达。由于所有引起FAM83H突变的A-均为终止在最后一个编码外显子（外显子5）中终止翻译的废话突变（外显子5），因此我们进一步假设FAM83H与其他细胞蛋白相互作用以形成功能复合物。我们提出了四个特定的目的：SA 1：表征牙肠发生过程中FAM83H表达的时间和空间模式并确定其亚细胞定位。 SA 2：分离FAM83H蛋白的结构和功能表征。 SA 3：识别FAM83H相互作用的蛋白质。 SA 4：确定截短的FAM83H的表达是否会干扰关节作用。方法：牙生成期间FAM83H表达的时间和空间模式由原位杂交和免疫组织化学确定。重组和本地FAM83H用于结构和功能研究。 FAM83H相互作用的蛋白质是使用酵母两杂交系统鉴定的，并使用哺乳动物的两杂化系统，共同沉淀和远西方分析进行了验证。正常和突变的FAM83H以成木细胞为转基因及其对牙釉质形成的影响。荒地相关性：具有遗传搪瓷缺陷（AI）患者的牙齿疼痛，较低的牙齿，自尊心较低，并认为自己具有劣质的生活质量。 FAM83H对于牙齿牙釉质形成至关重要的发现是我们对AI病因的理解。拟议的研究可能会导致我们对成熟细胞中重要活动的理解以及AI的新候选基因的突破。 公共卫生相关性：我们的小组确定FAM83H（具有序列相似性的家族83成员h）的缺陷在8号染色体上会导致严重的搪瓷畸形（常染色体显性低钙化的低钙化amelelogenogeny Imperfecta）。我们的目标是发现FAM83H的正常功能以及该蛋白质中的缺陷如何导致搪瓷畸形。我们的长期目标是提高对正常和病理牙齿形成的理解，以刺激公众口腔健康的改善。