Transcriptional regulation of liver specification in Xenopus tropicalis

热带爪蟾肝脏规格的转录调控

• 批准号: 8119663
• 负责人: Andrea Elizabeth Wills
• 金额: $ 5.13万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8119663
• 关键词: Address; Anterior; Antibodies; Basic Science; Binding; Biochemical; Biological Models; Cardiac; Cells; Collaborations; Commit; Communities; Competence; Cues; Databases; Development; Diagnosis; Dose; Embryo; Endoderm; Expressed Sequence Tags; FGFR4 gene; Fibroblast Growth Factor; Fostering; Gene Expression; Gene Targeting; Genes; Genetic; Goals; Hepatic; Hepatocyte; In Situ Hybridization; Injection of therapeutic agent; Investigation; Length; Link; Liver; Liver diseases; Mediating; Mesoderm; Methodology; Mus; Neurula; Nodal; Oligonucleotides; Organ; Organogenesis; Orthologous Gene; Pattern; Primitive foregut structure; Process; Program Development; Proteins; Protocols documentation; Rana; Regulation; Research; Screening procedure; Signal Transduction; Signaling Molecule; Specific qualifier value; Staging; Structure of septum transversum; Testing; Therapeutic; Time; Training; Transcriptional Regulation; Transplant Recipients; Western Blotting; Xenopus; cell motility; cell type; chromatin immunoprecipitation; embryonic stem cell; extracellular; human embryonic stem cell; in vivo; inhibitor/antagonist; insight; liver transplantation; loss of function; novel; overexpression; precursor cell; public health relevance; transcription factor

DESCRIPTION (provided by applicant): The establishment of a committed cell fate from a pluripotent precursor cell requires the coordinated integration of cell autonomous and non-autonomous cues through time. In the case of the liver, hepatic cell fate is thought to be acquired through a sequential process of commitment: cells are first directed to an endoderm fate by extracellular Nodal signaling, then acquire liver competence through the expression of specific transcription factors such as Gata4 and FoxA1, then are restricted from developing into other anterior endoderm organs by the action of extracellular signals including BMP signals from the septum transversum and FGF signals from the cardiac mesoderm. The goal of this research is to answer three unresolved questions in liver fate specification: which of the direct targets of Nodal signaling act in establishing liver competence; what are the direct targets of FoxA1; and what is the in vivo mechanism by which BMP and FGF regulate hepatic induction? These questions address gaps in our understanding of the transcriptional hierarchy through which known factors in liver development act, and of how each stage of liver development is mechanistically linked to the next. Answering these questions directly requires large numbers of embryos at early developmental stages, a limitation of amniote model systems where many previous studies on liver development have been conducted. Therefore, the proposed research will be carried out in the frog Xenopus tropicalis, which has large numbers of readily manipulated embryos ideal for rapid screening and biochemical investigations. The proposed project will generate a database of in situ hybridization expression patterns representing Nodal signaling targets, which will be made available to the community. The proposed project will also entail training and the development of expertise in recently-developed biochemical and high-throughput sequencing methodologies, specifically including the adaptation of existing protocols for chromatin immunoprecipitation and high throughput sequencing (ChIP-Seq) for use in X. tropicalis embryos with FoxA1 antibodies. Finally, this project will foster collaborations between basic research carried out in X. tropicalis and applications of the findings to directed differentiation of human embryonic stem cells to liver fates. The answers to these questions will elaborate our understanding of liver organogenesis, and will have applications to the development of new protocols for deriving hepatocytes from embryonic stem cells, a major goal for the eventual treatment of liver disease. PUBLIC HEALTH RELEVANCE: Understanding the steps that occur as a cell changes from a pluripotent fate to a liver fate during normal embryological development has and will continue to directly inform efforts to control the differentiation of human embryonic stem cells to liver cells. Liver cells derived from embryonic stem cells or other non-liver cell types are a potential therapeutic alternative to liver transplant for patients suffering from liver diseases. Further, a mechanistic understanding of the stages of liver development will provide insight into the genetic or developmental origins of some liver diseases arising from improper execution of the liver development program, and how they could be diagnosed or treated.

描述（由申请人提供）：从多能前体细胞建立定向细胞命运需要细胞自主和非自主线索随时间的协调整合。在肝脏的情况下，肝细胞命运被认为是通过连续的承诺过程获得的：细胞首先通过细胞外Nodal信号传导被引导至内胚层命运，然后通过表达特异性转录因子如Gata 4和FoxA 1获得肝能力，然后通过细胞外信号（包括来自隔膜的BMP信号）的作用限制其发育成其它前内胚层器官来自心脏中胚层的横突和FGF信号。本研究的目的是回答三个悬而未决的问题，在肝脏命运的规范：Nodal信号的直接目标行为在建立肝脏能力;什么是FoxA 1的直接目标;和什么是BMP和FGF调节肝诱导的体内机制？这些问题解决了我们对肝脏发育中已知因子作用的转录层次的理解中的差距，以及肝脏发育的每个阶段如何与下一个阶段机械地联系在一起。直接解决这些问题需要大量处于早期发育阶段的胚胎，这是许多先前关于肝脏发育的研究已经进行的胚胎移植模型系统的局限性。因此，拟议的研究将在热带爪蟾蛙中进行，热带爪蟾蛙具有大量易于操作的胚胎，非常适合快速筛选和生化研究。拟议的项目将产生一个代表Nodal信号靶标的原位杂交表达模式数据库，该数据库将提供给社区。拟议的项目还将需要在最近开发的生化和高通量测序方法方面进行培训和开发专业知识，特别是包括调整现有的染色质免疫沉淀和高通量测序（ChIP-Seq）方案以用于X。FoxA 1抗体。最后，该项目将促进在X开展的基础研究之间的合作。Tropicalis和应用的研究结果，以指导分化的人胚胎干细胞的肝脏命运。这些问题的答案将阐述我们对肝脏器官发生的理解，并将应用于开发从胚胎干细胞获得肝细胞的新方案，这是最终治疗肝脏疾病的主要目标。 公共卫生相关性：了解在正常胚胎发育过程中细胞从多能性命运转变为肝命运所发生的步骤已经并将继续直接告知控制人类胚胎干细胞分化为肝细胞的努力。来自胚胎干细胞或其他非肝细胞类型的肝细胞是肝脏疾病患者肝移植的潜在治疗替代方案。此外，对肝脏发育阶段的机械理解将提供对由于肝脏发育计划执行不当而引起的一些肝脏疾病的遗传或发育起源的洞察，以及如何诊断或治疗它们。