Genetic and molecular mechanisms of Xbp-1 mediated salivary gland development and differentiation

Xbp-1介导唾液腺发育和分化的遗传和分子机制

• 批准号: 10678146
• 负责人: Theresa Wrynn
• 金额: $ 3.33万
• 依托单位: STATE UNIVERSITY OF NEW YORK AT BUFFALO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10678146
• 关键词: Ablation; Acetylation; Acinar Cell; Address; Adult; Antibodies; Area; Automobile Driving; Binding; Binding Proteins; Bioinformatics; Biological; Biological Assay; Biological Process; Biology; Cell Differentiation process; Cell Lineage; Cell Maintenance; Cell physiology; Cells; ChIP-seq; Complex; Critical Pathways; Data; Data Set; Development; Differentiated Gene; Differentiation Antigens; Disease; Embryo; Embryonic Development; Enhancers; Ensure; Equilibrium; Factor X; Functional disorder; Gene Expression; Genes; Genetic; Genetic Transcription; Genomics; Gland; Goals; Histologic; Histones; Homeostasis; Human; Knockout Mice; Knowledge; Lysine; Maintenance; Maps; Measures; Mediating; Mediator; Mining; Molecular; Morphogenesis; Morphology; Mus; Muscarinic Acetylcholine Receptor; Natural regeneration; Neonatal; Normal tissue morphology; Organogenesis; Outcome; Pathway interactions; Patients; Pattern; Phenotype; Play; Polymerase Chain Reaction; Population; Process; Proliferating; Protein Isoforms; Proteins; Publishing; Quantitative Reverse Transcriptase PCR; RNA Splicing; Regulator Genes; Regulatory Element; Role; Salivary; Salivary Gland Diseases; Salivary Glands; Signal Pathway; Signal Transduction; Structure; Submandibular gland; Systems Biology; Testing; Therapeutic; Tight Junctions; Time; Tissues; XBP1 gene; Xerostomia; cell type; chromatin immunoprecipitation; conditional knockout; epigenomics; experimental study; gene regulatory network; genetic approach; gland development; improved; in vivo; in vivo Model; insight; interest; knockout animal; mouse model; polarized cell; postnatal; programs; regeneration following injury; response; saliva mediated; saliva secretion; single-cell RNA sequencing; spatiotemporal; therapeutic target; tissue regeneration; transcription factor; transcriptome sequencing; transcriptomics

Project Summary During embryogenesis and in adult, the delicate balance between proliferation and differentiation of the diverse cellular populations in the Salivary Gland (SG) must be tightly regulated to ensure normal tissue development and homeostasis. A better understanding of the regulatory mechanisms driving the cell fate decisions and lineage choices that anchor salivary gland morphogenesis will help identify pathways that are critical for tissue regeneration following injury, damage, or during diseased states. Xbp-1, specifically the spliced and transcriptionally active Xbp-1, isoform, has been implicated in Unfolded Protein Response (UPR), differentiation and a host of other biologically important cellular processes. Our knowledge of Xbp-1 in context of the SG is limited to scRNA-seq data showing Xbp-1 to be highly expressed in pro-acinar cells (the precursors to acinar cells) as well as a previously uncharacterized acinar cell phenotype observed in Xbp-1 null mice. The highly enriched expression of Xbp-1 in the SG and its association with super-enhancers as revealed by our analysis of genomic and epigenomics datasets, has led us to hypothesize that Xbp-1 plays an important and molecularly deterministic role in regulating cellular identity and function in the SG. However, the molecular mechanisms through which Xbp-1, dictates cell fate decisions and controls specific gene expression programs during SG development remains unexplored. To address these knowledge gaps, we will utilize conditional Xbp-1 knockout mouse models and sophisticated genomic/transcriptomic approaches to study three independent areas of interest. First, we will identify the spatiotemporal expression pattern of Xbp-1 in diverse SG cell types and measure the relative abundance of unspliced and spliced isoforms through developmental stages and during adult gland maintenance utilizing immunostainings and qRT-PCR (Aim 1). Second, we will use a conditional knockout (cKO) mouse model to determine the role of Xbp-1, during SG development and adult homeostasis (Aim 2). Such studies are necessary as they will identify, for the first time, the in vivo functional role of Xbp-1 in the SG. Finally, we will perform mechanistic studies to identify critical target genes and pathways that are governed by Xbp-1 and determine how the loss of Xbp-1 alters the gene expression program of the SG. Importantly, by implementing scRNA-seq studies, we will evaluate effects of loss of Xbp-1 on cell fate trajectories specifically during acinar development and maturation (Aim 3). Collectively, these studies will further our understanding of the Xbp1-dependent transcriptional and cellular networks important for the biological function of the SG particularly as it pertains to secretory acinar cell differentiation. Advances in our understanding of the underlying mechanisms driving SG development can provide a paradigm for regeneration, identify potential therapeutic targets to increase salivary flow in patients suffering from hyposalivation, and better inform therapies to treat patients with developmental dysfunctions and diseases of the salivary gland.

项目摘要 在胚胎发生和成人中，多样的增殖与分化之间的微妙平衡 必须严格调节唾液腺（SG）中的细胞种群，以确保正常的组织发展 和稳态。更好地理解驱动细胞命运决策的调节机制和 锚定唾液腺形态发生的谱系选择将有助于识别对组织至关重要的途径 受伤，损害或患病状态期间的再生。 XBP-1，特别是剪接的， 转录活性XBP-1（同工型）与未折叠的蛋白质反应（UPR）有关，分化 以及许多其他重要的生物学细胞过程。我们在SG背景下对XBP-1的了解是 仅限于SCRNA-seq数据，显示XBP-1在pro-Acinar细胞中高度表达（腺泡的前体 细胞）以及在XBP-1无效小鼠中观察到的先前未表征的腺泡细胞表型。高度 XBP-1在SG中的富集表达及其与超级增强剂的关联，我们的分析揭示了 基因组和表观基因组学数据集使我们假设XBP-1在重要且分子上发挥作用 确定性的作用在调节SG中的细胞身份和功能中。但是，分子机制 通过哪些XBP-1决定细胞命运决策并控制SG期间特定的基因表达程序 开发仍未开发。为了解决这些知识差距，我们将利用有条件的XBP-1淘汰赛 小鼠模型和复杂的基因组/转录组方法研究三个独立领域 兴趣。首先，我们将确定XBP-1在不同的SG细胞类型中的时空表达模式和 通过发育阶段和期间测量未植物和剪接的同工型的相对丰度 利用免疫染色和QRT-PCR的成年腺维持（AIM 1）。第二，我们将使用有条件的 敲除（CKO）小鼠模型以确定XBP-1的作用，在SG发育和成人稳态中 （目标2）。这样的研究是必要的，因为它们将首次确定XBP-1在体内功能作用 SG。最后，我们将进行机械研究，以识别关键的靶基因和途径 由XBP-1管辖，并确定XBP-1损失如何改变SG的基因表达程序。 重要的是，通过实施SCRNA-Seq研究，我们将评估XBP-1损失对细胞命运轨迹的影响 特别是在腺泡发展和成熟期间（AIM 3）。总的来说，这些研究将进一步 了解XBP1依赖性转录和细胞网络对生物功能很重要 SG特别是与分泌的腺泡细胞分化有关。我们对理解的进步 驱动SG开发的基本机制可以为再生提供范式，确定潜力 治疗靶点以增加缺血患者的唾液流量，并更好地告知疗法 治疗唾液腺发育功能障碍和疾病的患者。