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

• 批准号: 10471183
• 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
• 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/10471183
• 关键词: 3-Dimensional; Adhesions; Affect; American; Architecture; Autoimmune Diseases; B cell differentiation; Beta Cell; Binding; Bioinformatics; Cell Differentiation process; Cell Nucleus; Cell Polarity; Cell Proliferation; Cell model; Cells; Clinical Trials; Cultured Cells; Cytoplasm; Data; Defect; Development; Diabetes Mellitus; Embryo; Epithelial; Epithelial Cell Proliferation; Epithelial Cells; Evaluation; Failure; Gene Expression Profiling; Generations; Genes; Genetic; Goals; Homeostasis; In Vitro; Insulin; Insulin-Dependent Diabetes Mellitus; Islet Cell; Islets of Langerhans; Maintenance; Molecular; Morphogenesis; Mosaicism; Multipotent Stem Cells; Mus; Natural regeneration; Organ; Organ Size; Organoids; Pancreas; Pancreatic Bud; Pathway interactions; Pharmacology; Phosphotransferases; Population; Process; Regulation; Replacement Therapy; Role; Shapes; Signal Pathway; Signal Transduction; Testing; Therapeutic; Tissue Culture Techniques; Transcription Coactivator; Transgenic Mice; Transitional Cell; 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; progenitor; regenerative therapy; stem cells; transcription factor; transcriptome sequencing; translational impact

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3000万美国人患有前或现有的糖尿病。治疗糖尿病的一种创新方法是产生功能性β细胞，该β细胞起源于早期胰腺芽中的多能祖细胞种群。这种方法目前正在临床试验中，但公认的问题是次优祖细胞产生。尽管稍后胰腺发育的机制已得到很好的定义，但胰腺祖细胞规范的调节尚不清楚。该建议将使用转基因小鼠模型以及包括器官外植体和胰腺器官的组织培养技术检查河马信号通路下游的遗传途径。生物信息学方法将用于（1）确定河马信号传导是否调节胰腺祖细胞规范，形态发生或细胞分化，以及（2）确定调节YAP1/TAZ和NFκB的串扰机制，该机制调节了pancreas中祖细胞规范和BetA细胞分化之间的YAP1/TAZ和NFκB。我们发现，删除LATS激酶会失调上皮细胞增殖，从而导致异常形态发生和细胞分化，从而支持胰腺形态发生与细胞分化紧密相关的观念。使用转录分析，我们发现YAP1/TAZ促进了NFκB激活因子基因，在这里我们将定义所涉及的机制。在此提案中，我们提出了一个独特的想法，即河马途径成分充当控制在此过程中NFκB活性水平的变阻器，从而雕刻了产生β细胞的上皮细胞。该利基是胚胎胰腺核心的专业和短暂上皮丛。我们假设需要LATS1/2活性是需要体内稳态才能使YAP1/TAZ灭活，从而抑制了正常胰腺中的异常NFκB信号传导。胰腺上皮的NFκB活性升高会导致EMT倡议失调和β细胞命运的丧失，这是由于上皮丛生的破坏而导致的。我们假设升高的YAP1活性至少部分通过pantetheinase vanin1（VNN1）刺激NFκB信号传导。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：1）检查LATS1和LATS2的丢失如何影响上皮完整性以及上皮到间质转变（EMTS）的启动； 2）检查YAP1的过表达是否模仿单个或簇上皮细胞中的LATS1/2双重缺失（1/2DKO），以及对在1/2DKO中已知的下游靶标的评估； 3）评估NFKB途径在胰腺上皮稳态中的作用，并检查VNN1在此过程中的作用。我们希望这项工作能够增强下游靶标在胰腺祖细胞和β细胞中的转化影响，以热β细胞的替代和再生以治疗糖尿病。