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.

项目摘要/摘要 上牙可能会引起广泛的牙齿​​并发症。随着额外的牙齿形成 现有的牙列，阐明了超生物牙齿形成的分子机制不仅有助于 制定该疾病的治疗策略，但也为牙齿再生提供了见解。尽管 理解形态因素，生长因素和 在超级牙齿形成中的转录因子，对细胞外的作用知之甚少 在此病理过程中，诸如蛋白聚糖之类的成分。我们最近的研究表明，失活 牙齿上皮FAM20B，这是一种新发现的木糖激酶，对糖胺聚糖（GAG）必不可少 组装，导致小鼠的超努切牙。我们的试点研究表明， 牙齿上皮导致Wnt信号的异位激活，并且异位SOX2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 表明在某些FAM20B催化的蛋白聚糖上的插科打d抑制了Wnt信号，但有助于明智 介导的抑制Wnt。相反，将Wnt抑制剂施用到突变胚胎中 在某些情况下，牙齿表型。这些数据导致我们提出了某些FAM20B-的中心假设 通过阴性调节介导干细胞更新，催化蛋白聚糖调节牙齿更新 在牙齿上皮中的Wnt信号传导上。为了检验这一假设，我们提出以下三个特定 目的：（1）确定FAM20B催化的蛋白聚糖是否通过阴性调节介导牙齿更新 在Wnt信号传导上，如果GAG介导的WISE对Wnt的明智抑制是超生物牙齿的基础 形成。我们将通过过表达DKK1或明智的牙齿上皮来执行“救援”实验 抑制K14CRE/+; FAM20BFL/FL小鼠中过度活化的Wnt信号传导。 （2）确定FAM20B-是否 催化的蛋白聚糖通过介导牙齿上皮细胞更新来调节牙齿更新。 我们将执行谱系跟踪和诱导敲除实验，以确定干细胞的作用 续签缺乏症引起的超级牙齿，如果FAM20B催化的PG介导了茎 细胞上皮的细胞更新以细胞自主方式。 （3）识别FAM20B催化 蛋白聚糖造成超级牙齿形成。我们将确定表达方式 牙齿中FAM20B催化的蛋白聚糖，并确定那些对Wnt信号传导的负面调节的蛋白聚糖。这 候选蛋白聚糖的生物学功能将由基因敲低和器官培养确定 方法。这项研究的完成将提高我们对分子机制的理解 底部的牙齿形成，并有助于为牙齿再生奠定基础。作为 FAM20B和蛋白聚糖在许多组织中广泛表达，从中获得的知识 研究还可以提供对其他组织中蛋白聚糖介导的信号传导的宝贵见解。