The Role of FAM20B-catalyzed Proteoglycans in Tooth Development

FAM20B 催化蛋白多糖在牙齿发育中的作用

• 批准号: 9899242
• 负责人: XIAOFANG WANG
• 金额: $ 35.27万
• 依托单位: TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9899242
• 关键词: Ablation; Address; Area; Biological Process; Cell surface; Cells; Data; Dental; Dentition; Development; Disease; Dose; Embryo; Enamel Organ; Epithelial; Epithelium; Equilibrium; Etiology; Extracellular Matrix; Family; Genes; Glycosaminoglycans; Growth Factor; Human; In Vitro; Incisor; Knock-out; Knowledge; Location; Mediating; Methods; Molecular; Mus; Organ Culture Techniques; Pathologic Processes; Pattern; Phenotype; Phosphotransferases; Pilot Projects; Play; Proteins; Proteoglycan; Regulation; Research; Rodent; Role; Side; Signal Transduction; Signaling Molecule; Site; Supernumerary Tooth; Testing; Therapeutic; Tissues; Tooth regeneration; Tooth structure; WNT Signaling Pathway; Xylose; adenoviral-mediated; design; epithelial stem cell; experimental study; extracellular; inhibitor/antagonist; insight; knock-down; member; morphogens; mutant; novel; overexpression; programs; stem cell division; stem cells; transcription factor

Project summary/Abstract Supernumerary teeth can cause a broad range of dental complications. As extra teeth are formed on existing dentition, unraveling the molecular mechanism of supernumerary tooth formation will not only help develop the therapeutic strategy for this disease but also provide insights into tooth regeneration. Despite the significant progress in understanding the regulatory role of morphogens, growth factors, and transcriptional factors in supernumerary tooth formation, little is known about the role of extracellular components such as proteoglycans in this pathological process. Our recent studies show that inactivation of dental epithelial Fam20B, a newly discovered xylose kinase essential for glycosaminoglycan (GAG) assembly, leads to supernumerary incisors in mice. Our pilot study reveals that GAG deficiency in the dental epithelium leads to ectopic activation of WNT signaling, and that an ectopic Sox2 expression is located in the same area, which normally should disappear from this site after E14.5. Our in vitro study shows that GAGs on certain FAM20B-catalyzed proteoglycans suppress WNT signaling but facilitate Wise- mediated inhibition on WNT. Conversely, administering WNT inhibitor to the mutant embryos rescued the tooth phenotype in some cases. These data led us to form our central hypothesis that certain FAM20B- catalyzed proteoglycans regulate tooth renewal by mediating the stem cell renewal via negative regulation on WNT signaling in the dental epithelium. To test this hypothesis, we propose the following three specific aims: (1) To determine if FAM20B-catalyzed proteoglycans mediate tooth renewal via negative regulation on WNT signaling, and if GAG-mediated Wise inhibition on WNT underlies the supernumerary tooth formation. We will perform "rescue" experiments by overexpressing DKK1 or Wise in the dental epithelium to inhibit the overactivated WNT signaling in K14Cre/+;Fam20Bfl/fl mice. (2) To determine whether FAM20B- catalyzed proteoglycans regulate tooth renewal by mediating the stem cell renewal in the dental epithelium. We will perform lineage tracing and inducible knockout experiments to determine the role of stem cell renewal in GAG deficiency-caused supernumerary teeth, and if the FAM20B-catalyzed PGs mediate stem cell renewal in the dental epithelium in a cell-autonomous manner. (3) To identify the FAM20B-catalyzed proteoglycans responsible for the supernumerary tooth formation. We will determine the expression pattern of FAM20B-catalyzed proteoglycans in tooth and identify those negatively regulating WNT signaling. The biological function of candidate proteoglycans will be determined by gene knockdown and organ culture methods. The completion of this study will advance our understanding about the molecular mechanism underlying supernumerary tooth formation and help in laying the groundwork for tooth regeneration. As FAM20B and proteoglycans are extensively expressed in many tissues, the knowledge gained from this study may also provide valuable insights into the proteoglycan-mediated signaling in other tissues.

项目概要/摘要 多生牙会引起一系列的牙科并发症。当额外的牙齿在 现有的牙列，解开多生牙形成的分子机制，不仅有助于 为这种疾病制定治疗策略，同时也为牙齿再生提供见解。尽管 在了解形态发生素、生长因子和 转录因子在多生牙形成中的作用，对细胞外转录因子的作用知之甚少。 蛋白多糖等成分参与了这一病理过程。我们最近的研究表明， 牙上皮Fam 20 B，一种新发现的糖胺聚糖（GAG）必需的木糖激酶 组装，导致小鼠多出门牙。我们的初步研究表明，GAG缺乏症， 牙齿上皮导致WNT信号的异位激活，并且异位Sox 2表达是 位于同一区域，通常应在E14.5之后从该站点消失。我们的体外研究 显示某些FAM 20 B催化的蛋白聚糖上的GAG抑制WNT信号传导，但促进WISE- 介导对WNT的抑制。相反，给突变胚胎施用WNT抑制剂， 在某些情况下的牙齿表型。这些数据使我们形成了我们的中心假设，即某些FAM 20 B- 催化的蛋白多糖通过负调节介导干细胞更新来调节牙齿更新 对牙齿上皮中WNT信号的影响。为了验证这一假设，我们提出了以下三个具体的 目的：（1）研究FAM 20 B催化的蛋白多糖是否通过负调控介导牙齿更新 如果GAG介导的WNT抑制是多生牙的基础， 阵我们将通过在牙上皮中过表达DKK 1或Wise来进行“拯救”实验， 抑制K14 Cre/+; Fam 20 Bfl/fl小鼠中过度激活的WNT信号传导。(2)为了确定FAM 20 B是否- 催化的蛋白聚糖通过介导牙上皮中的干细胞更新来调节牙齿更新。 我们将进行谱系追踪和诱导性基因敲除实验，以确定干细胞的作用， GAG缺乏引起的多生牙的更新，如果FAM 20 B催化的PG介导干 以细胞自主的方式在牙上皮中进行细胞更新。(3)为了鉴定FAM 20 B催化的 蛋白多糖负责多生牙的形成。我们将确定表达模式 FAM 20 B催化的蛋白聚糖在牙齿和鉴定负调节WNT信号。的 候选蛋白聚糖的生物学功能将通过基因敲除和器官培养来确定 方法.本研究的完成将进一步加深我们对分子机制的理解 潜在的多生牙的形成和帮助奠定基础的牙齿再生。作为 FAM 20 B和蛋白聚糖在许多组织中广泛表达，由此获得的知识 这项研究也可能对其他组织中蛋白聚糖介导的信号传导提供有价值的见解。