Elucidating the role of Polycomb Repressive Complexes in Lingual Papillae Development

阐明多梳抑制复合物在舌乳头发育中的作用

• 批准号: 10424544
• 负责人: Elena Ezhkova
• 金额: $ 35.83万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10424544
• 关键词: Address; Anterior; Antibodies; Basal Cell; Binding; Biological Assay; Candidate Disease Gene; Cell Differentiation process; Cell Extracts; Cell Lineage; Cell Maintenance; Cells; ChIP-seq; Chromatin; Communication; Complex; Connective Tissue; Coupled; Data; Defect; Development; Drosophila genus; Eating; Embryo; Embryonic Development; Epithelial; Epithelial Cells; Esthesia; Failure; Filiform Papilla; Fungiform Papilla; Gene Expression; Genes; Genetic; Genetic Transcription; Goals; Histone H2A; In Vitro; Ligands; Maintenance; Mastication; Mediating; Mesenchymal; Microglossias; Molecular; Morphogenesis; Mus; Muscle; Ontology; Organ; PRC1 Protein; Pathway interactions; Pattern; Pattern Formation; Phenotype; Play; Polycomb; Process; Protocols documentation; RNA; Regulation; Research Proposals; Role; SHH gene; Side; Signal Pathway; Structure; Taste Buds; Taste Perception; Testing; Tissues; To specify; Tongue; Transcription Repressor; Transcriptional Regulation; WNT Signaling Pathway; appendage; base; chromatin immunoprecipitation; embryonic stem cell; experimental study; gene repression; in vivo; insight; keratinization; loss of function; novel; oral tissue; progenitor; recruit; smoothened signaling pathway; stem cells; tongue papilla; transcription factor; transcriptome sequencing

Summary The tongue is a complex organ that is essential for mastication, taste sensation, and communication. Proper development of the tongue is dependent on complex interactions between different tissues, including muscle, connective tissue, and epithelium. Importantly, failure to establish these interactions leads to developmental defects, such as aglossia, microglossia, and ankyloglossia. The lingual epithelium is an array of epithelial-mesenchymal appendages termed papillae. Fungiform papillae harbor taste buds required for taste sensation. They are surrounded by keratinized, non-gustatory filiform papillae, which provide essential barrier functions. Wnt and Shh signaling pathways are central players in regulating the patterning and development of fungiform and filiform papillae; however, the role of cell-intrinsic mechanisms in the specification and maintenance of these structures remains unclear. Polycomb repressive complex (PRC) 1 is a major chromatin transcriptional regulator and has been shown to play a critical role in lineage identity and early embryonic development; however, PRC1-mediated regulation of cell lineages has yet to be explored in oral tissues. To investigate the role of PRC1 in the development of lingual papillae, we generated mice in which essential PRC1 subunits, Ring1a and Ring1b, were conditionally ablated in the basal lingual epithelial cells (Ring1a/b 2KO). Our results showed that in Ring1a/b 2KO mice, the fungiform papillae are not maintained and the filiform papillae are not formed. Further analysis of the Ring1a/b-null lingual epithelium revealed widespread expression of the Sonic Hedgehog (Shh) ligand and the transcription factor Sox2, whereas in control mice, the expression of these genes was confined to taste progenitors. The expression pattern of components of the Wnt signaling pathway, Wnt10b and Lef1, was also altered in Ring1a/b 2KO; however, this did not result in aberrant canonical Wnt signaling. We hypothesize that PRC1 restricts gene expression and thus promotes lingual papilla patterning and formation. To dissect the molecular mechanisms of PRC1's control of the lingual epithelium, we will carry out an RNA-seq analysis of FACS-purified control and Ring1a/b- null lingual epithelial cells. This analysis will be coupled with ChIP-seq with antibodies against PRC1 subunits to identify the direct targets of PRC1. To identify the specific molecular pathways disrupted in the Ring1a/b-null lingual epithelium, we will carry out Gene Ontology analysis and focus on candidate genes that regulate epithelial-mesenchymal interactions, transcriptional regulation, and lineage identity. We will analyze the functional significance of the candidate genes by performing genetic experiments in vivo. Together, our studies will allow us to elucidate PRC1's regulation of the lingual epithelium, including its interaction with developmental signaling pathways. Understanding the complex molecular interactions between lingual tissues may also help us to reveal the mechanisms that cause congenital tongue abnormalities to form.

摘要 舌头是一个复杂的器官，对咀嚼、味觉和交流是必不可少的。 舌头的正常发育依赖于不同组织之间的复杂相互作用，包括 肌肉、结缔组织和上皮组织。重要的是，未能建立这些互动会导致 发育性缺陷，如失语症、小舌症和舌强症。舌上皮由一系列 上皮间充质附属物，称为乳头。菌状乳头含有味觉所需的味蕾 轰动一时。它们周围是角化的、非味觉的丝状乳头，构成了基本的屏障。 功能。WNT和Shh信号通路在调节细胞的形成和发展中起着核心作用。 菌状和丝状乳头；然而，细胞内在机制在规范和 这些建筑的维护情况仍不明朗。多梳抑制复合体(PRC)1是一种主要的染色质 转录调节因子，已被证明在血统认同和早期胚胎中起着关键作用 然而，在口腔组织中，Prc1介导的细胞谱系的调节尚未被探索。 为了研究Prc1在舌乳头发育中的作用，我们建立了 基本的PRC1亚基Ring1a和Ring1b在基本的舌上皮细胞中被有条件地消融 (Ring1a/b 2KO)。我们的结果表明，在Ring1a/b 2KO小鼠中，菌状乳头不被保留和 未形成丝状乳突。进一步分析Ring1a/b缺失的舌上皮 广泛表达Sonic Hedgehog(Shh)配体和转录因子Sox2，而在 对照小鼠，这些基因的表达仅限于味觉祖细胞。的表达方式 在Ring1a/b 2KO中，Wnt信号通路的组成部分Wnt10b和Lef1也发生了改变；然而，这 没有导致异常的规范的Wnt信号。我们假设Prc1限制了基因的表达 从而促进舌乳头的图案化和形成。PRc1‘S发病的分子机制剖析 为了控制舌上皮，我们将对FACS纯化对照和Ring1a/b-Ring1a-b进行RNA-SEQ分析。 舌上皮细胞为空。这一分析将与带有Prc1亚单位抗体的CHIP-SEQ相结合 确定PRC1的直接靶点。识别Ring1a/b中被破坏的特定分子通路-空 舌上皮，我们将进行基因本体论分析，并重点关注调控的候选基因 上皮-间充质的相互作用，转录调控，和谱系认同。我们将分析 通过在体内进行遗传学实验来研究候选基因的功能意义。 总之，我们的研究将使我们能够阐明Prc1‘S对舌上皮的调控，包括它的 与发育信号通路的相互作用。理解分子间复杂的相互作用 舌组织也可能帮助我们揭示导致先天性舌部畸形形成的机制。