Chromatin modifier Polycomb Repressive Complex 2 as a regulator of dental epithelial progenitor cells

项目摘要/摘要 了解牙齿上皮祖细胞命运决定的机制将有助于为长期 干细胞生物学在人类牙列中的临床应用基础。在不断生长的小鼠中 切牙，周期祖细胞在动态平衡和损伤期间协调切牙上皮的更新- 诱导再生。祖细胞自我更新和分化的迅速和协调的平衡 无论是形成釉质的成釉细胞还是非成釉细胞，都能支持牙齿的持续生长。 然而，控制这些快速命运决定的机制仍不清楚。要了解机制 关于细胞命运的决定，这项提议将研究多梳抑制复合体2(PRC2)是如何通过 组蛋白3上赖氨酸27的三甲基化，抑制基因表达以驱动祖细胞的命运承诺。 这项建议将确定PRC2催化亚基增强子Zust Homolog 2(EZH2)在切牙中的作用 上皮细胞及其在祖细胞分化过程中的特定染色质靶点。目标1将研究EZH2如何 在动态平衡和损伤诱导过程中对切牙上皮祖细胞命运的影响 再生。这将通过确定祖细胞中Ezh2的丢失如何影响细胞命运决定来实现， 如分化、自我更新或细胞凋亡的变化。目标2将阐明EZH2的特定靶点 在动态平衡和损伤诱导再生期间的祖细胞中。切牙的染色质状态 这两种情况下的上皮细胞将使用最先进的转座酶单细胞测定法来确定 可及染色质测序(ScATACseq)和靶下切割及使用核酸酶释放 (削减和运行)技术。这些分析将显示切牙上皮中的PRC2靶点是否相似 在其他自我更新的组织中。他们还将确定牙齿上皮组织特异性靶点 对成釉细胞和非成釉细胞命运有贡献。 这项研究计划将与旨在制定 申请者的职业是牙医-科学家。培训包括来自高资质人员的结构化指导 临床医生-科学家赞助，以及通过参加研讨会、期刊俱乐部、 课堂、实验室会议、会议等。研究和培训将在华盛顿大学进行 加州，旧金山，它提供了一流的研究环境和一所优秀的牙科学校 用于临床培训。

Project Summary/Abstract Understanding the mechanisms of dental epithelial progenitor cell fate decisions will help to lay the long-term groundwork for clinical applications of stem cell biology in human dentition. In the continuously growing mouse incisor, cycling progenitor cells orchestrate incisor epithelium renewal during both homeostasis and injury- induced regeneration. The swift and well-orchestrated balance of progenitor cell self-renewal and differentiation into either enamel-producing ameloblasts or non-ameloblasts supports the continuous growth of the tooth. However, the mechanisms that control these rapid fate decisions remain unclear. To understand mechanisms of cell fate decisions, this proposal will examine how Polycomb Repressive Complex 2 (PRC2), through trimethylation of lysine 27 on Histone 3, represses gene expression to drive fate commitment of progenitor cells. This proposal will establish the role of PRC2 catalytic subunit Enhancer of Zeste Homolog 2 (EZH2) in the incisor epithelium and its specific chromatin targets during progenitor differentiation. Aim 1 will examine how EZH2 contributes to incisor epithelial progenitor cell fate decisions during homeostasis and injury-induced regeneration. This will be achieved by determining how loss of Ezh2 in progenitor cells affects cell fate decisions, such as changes to differentiation, self-renewal or apoptosis. Aim 2 will elucidate the specific targets of EZH2 in progenitor cells during homeostasis and injury-induced regeneration. Chromatin states of incisor epithelial cells during both conditions will be determined using state-of-the-art single-cell Assay for Transposase Accessible Chromatin sequencing (scATACseq) and Cleavage Under Targets and Release Using Nuclease (CUT&RUN) technologies. These analyses will show whether PRC2 targets in the incisor epithelium are similar to those in other self-renewing tissues. They will also identify the dental epithelial tissue-specific targets that contribute to ameloblast and non-ameloblast fates. This research plan will be conducted in conjunction with a comprehensive training plan designed to develop the applicant’s career as a dentist-scientist. The training includes structured mentorship from a highly qualified clinician-scientist sponsor, as well as scientific and technical training through attending seminars, journal clubs, classes, laboratory meetings, conferences, and more. Research and training will take place at the University of California, San Francisco, which offers both an outstanding research environment and an excellent dental school for clinical training.