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

基本信息

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
项目类别：
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

项目摘要

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.

项目概要在胚胎发生和成体过程中，不同物种的增殖和分化之间存在微妙的平衡。必须严格调节唾液腺 (SG) 中的细胞群，以确保正常的组织发育和体内平衡。更好地了解驱动细胞命运决定的调控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、损伤后或患病状态下的再生。 Xbp-1，特别是拼接和转录活性 Xbp-1（异构体）与未折叠蛋白反应 (UPR)、分化有关以及许多其他生物学上重要的细胞过程。我们对 SG 背景下的 Xbp-1 的了解是仅限于 scRNA-seq 数据显示 Xbp-1 在原腺泡细胞（腺泡细胞的前体）中高度表达细胞）以及在 Xbp-1 缺失小鼠中观察到的先前未表征的腺泡细胞表型。高度我们的分析揭示了 SG 中 Xbp-1 的富集表达及其与超级增强子的关联基因组和表观基因组学数据集，使我们假设 Xbp-1 在分子水平上发挥着重要作用 SG 中调节细胞身份和功能的确定性作用。然而，分子机制通过 Xbp-1，决定细胞命运决定并控制 SG 期间的特定基因表达程序发展仍有待探索。为了解决这些知识差距，我们将利用条件 Xbp-1 基因敲除小鼠模型和复杂的基因组/转录组方法来研究三个独立的领域兴趣。首先，我们将确定 Xbp-1 在不同 SG 细胞类型中的时空表达模式，测量发育阶段和发育过程中未剪接和剪接异构体的相对丰度利用免疫染色和 qRT-PCR 进行成人腺体维护（目标 1）。其次，我们将使用条件敲除 (cKO) 小鼠模型以确定 Xbp-1 在 SG 发育和成年体内平衡过程中的作用（目标 2）。此类研究是必要的，因为它们将首次确定 Xbp-1 在体内的功能作用 SG。最后，我们将进行机制研究，以确定关键的靶基因和途径由 Xbp-1 控制并确定 Xbp-1 的丢失如何改变 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 丢失对细胞命运轨迹的影响特别是在腺泡发育和成熟期间（目标 3）。总的来说，这些研究将进一步推动我们了解 Xbp1 依赖性转录和细胞网络对于生物功能很重要 SG，特别是因为它与分泌性腺泡细胞分化有关。我们对事物理解的进步驱动 SG 发展的潜在机制可以提供再生范例，识别潜力治疗目标是增加唾液分泌不足患者的唾液流量，并更好地为治疗提供信息治疗患有发育障碍和唾液腺疾病的患者。