Hippo suppression of NFkB controls pancreas morphogenesis and beta cell fate

Hippo 抑制 NFkB 控制胰腺形态发生和 β 细胞命运

基本信息

批准号：
10665660
负责人：
Ondine B Cleaver
金额：
$ 40.5万
依托单位：
UT SOUTHWESTERN MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-09-11 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10665660
关键词：
3-Dimensional Ablation Adhesions Affect American Architecture Autoimmune Diseases Beta Cell Binding Bioinformatics Cell Differentiation process Cell Nucleus Cell Polarity Cell Proliferation Cells Clinical Trials Cultured Cells Cytoplasm Data Defect Development Diabetes Mellitus Embryo Epithelial Cell Proliferation Epithelial Cells Epithelium Evaluation Failure Gene Expression Profiling Generations Genes Genetic Goals Homeostasis In Vitro Insulin Insulin-Dependent Diabetes Mellitus Intercellular Junctions Islet Cell Islets of Langerhans Maintenance Modeling Molecular Morphogenesis Mosaicism Multipotent Stem Cells Mus NF-kappa B Natural regeneration Organ Organ Size Organoids Pancreas Pancreatic Bud Pathway interactions Phosphorylation Phosphotransferases Population Process Proliferating Regulation Replacement Therapy Role Shapes Signal Pathway Signal Transduction Specific qualifier value Testing Therapeutic Tissue Culture Techniques Transcription Coactivator Transgenic Mice Vimentin Work beta cell replacement diabetes mellitus therapy epithelial to mesenchymal transition improved in vitro Model in vivo inhibitor innovation insight islet stem cells mouse model mutant novel overexpression pancreas development pantetheinase pharmacologic progenitor regenerative therapy stem cells transcription factor transcriptome sequencing translational impact

项目摘要

More than 30 million Americans have pre- or existing diabetes. An innovative approach to treat diabetes is to generate functional beta cells, which originate from a multipotent progenitor population in the early pancreas bud. This approach is currently in clinical trials, but a recognized problem is suboptimal progenitor generation. While mechanisms regulating later pancreas development have been well defined, how pancreas progenitor specification is regulated remains unclear. This proposal will examine genetic pathways downstream of the Hippo signaling pathway using transgenic mouse models, as well as tissue culture techniques that include organ explants and pancreatic organoids. Bioinformatic approaches will be used to (1) determine if Hippo signaling regulates pancreatic progenitor specification, morphogenesis, or cell differentiation, and (2) determine crosstalk mechanisms between Yap1/Taz and NFκB that regulate progenitor specification and beta cell differentiation in the pancreas. We have found that deleting the Lats kinases dysregulates epithelial cell proliferation, leading to aberrant morphogenesis and cell differentiation, supporting the idea that pancreatic morphogenesis is closely tied to cell differentiation. Using transcriptional profiling, we have found that Yap1/Taz promote NFκB activator genes and here we will define the mechanisms involved. In this proposal, we propose the unique idea that Hippo pathway components act as a rheostat to control levels of NFκB activity during this process, sculpting the epithelial niche that generates beta cells. This niche is a specialized and transient epithelial plexus at the core of the embryonic pancreas. We hypothesize that Lats1/2 activity is required homeostatically to inactivate Yap1/Taz and thereby suppress aberrant NFκB signaling in the normal pancreas. Elevated NFκB activity in pancreatic epithelium results in dysregulated EMT initiation and loss of β-cell fate, due to disruption of the epithelial plexus niche. We hypothesize that elevated Yap1 activity stimulates NFκB signaling at least in part via the pantetheinase Vanin1 (Vnn1). Our aims will use in vivo and in vitro models to investigate these observations. Our hypotheses will be tested via: 1) an examination of how loss of Lats1 and Lats2 affects epithelial integrity and initiation of epithelial-to-mesenchymal transitions (EMTs); 2) an examination of whether overexpression of Yap1 mimics the Lats1/2 double deletion (1/2DKO) in single or clusters of epithelial cells, as well as an assessment of downstream targets known to be affected in 1/2DKO; and 3) an evaluation of the role of the NFkB pathway in pancreatic epithelial homeostasis and an examination of the role of Vnn1 in this process. We hope this work will enhance translational impact of downstream targets in pancreas progenitors and beta cells, with the goal of therapeutic beta cell replacement and regeneration to treat diabetes.

超过 3000 万美国人患有糖尿病或现有糖尿病，治疗糖尿病的一种创新方法是产生功能性 β 细胞，这种细胞源自早期胰腺芽中的多能祖细胞群。这种方法目前正在进行临床试验，但这是一个公认的问题。虽然调节后期胰腺发育的机制已经明确，但如何调节胰腺祖细胞规范仍不清楚，该提案将使用转基因小鼠模型来检查 Hippo 信号通路下游的遗传途径。包括器官外植体和胰腺类器官的组织培养技术将用于 (1) 确定 Hippo 信号传导是否调节胰腺祖细胞规格、形态发生或细胞分化，以及 (2) 确定调节 Yap1/Taz 和 NFκB 之间的串扰机制。我们发现，删除 Lats 激酶会失调上皮细胞增殖，导致异常。使用转录分析，我们发现 Yap1/Taz 促进 NFκB 激活基因，支持胰腺形态发生与细胞分化密切相关的观点，在此我们将定义所涉及的机制。认为 Hippo 通路成分在此过程中充当变阻器来控制 NFκB 活性水平，塑造生成 β 细胞的上皮生态位。该生态位是一种特殊且短暂的上皮细胞。我们发现，Lats1/2 活性是稳态失活 Yap1/Taz 所必需的，从而抑制正常胰腺中异常的 NFκB 信号传导。胰腺上皮中 NFκB 活性升高导致 EMT 启动失调和 β 丧失。 -细胞命运，由于上皮神经丛生态位的破坏，我们发现 Yap1 活性升高会刺激 NFκB 信号传导。至少部分通过泛酸酶 Vanin1 (Vnn1) 我们的目标将使用体内和体外模型来研究这些观察结果，我们的假设将通过以下方式进行检验：1) 检查 Lats1 和 Lats2 的缺失如何影响上皮完整性和起始。上皮-间质转化 (EMT)；2) 检查 Yap1 的过表达是否模拟单或中的 Lats1/2 双缺失 (1/2DKO)上皮细胞簇，以及对已知在 1/2DKO 中受影响的下游靶标的评估；3) 评估 NFkB 通路在胰腺上皮稳态中的作用，并检查 Vnn1 在此过程中的作用。我们希望这项工作将增强胰腺祖细胞和β细胞下游靶标的转化影响，以实现治疗性β细胞替代和再生来治疗糖尿病。