Molecular mechanisms of cell fate specification

细胞命运规范的分子机制

基本信息

批准号：
7733927
负责人：
LYNNE M ANGERER
金额：
$ 107.47万
依托单位：
NATIONAL INSTITUTE OF DENTAL & CRANIOFACIAL RESEARCH
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7733927
关键词：
Achievement Agonist Andro-Diane Animals Bioinformatics Biological Assay Biological Models Cells Cilia Collaborations Development Developmental Biology Dopamine Dopamine Antagonists Eating Ecology Ectoderm Embryo Embryonic Development Endoderm Endomesoderm Event Evolution Food Forebrain Development G-Protein-Coupled Receptors Gene Components Gene Structure Genes Genetic Transcription Genome Germ Layers Growth Injection of therapeutic agent Intramural Research Program Introns Investigation Larva Lateral Length Manuscripts Mediating Mesoderm Methods Molecular Morphology Nerve Nervous system structure Neurons Non - mammalian blastula Numbers Operative Surgical Procedures Oral Pathway interactions Pattern Physiology Pigments Positioning Attribute Process Prosencephalon Refractory Regulator Genes Reporting Repression Research Rest Risk Role Sea Urchins Sensory Signal Pathway Signal Transduction Skeletal system Specific qualifier value Specificity Staging Structure System Tissues To specify Transforming Growth Factor beta Translations Upper arm Vertebrates Work animal tissue base blastocyst cell type density dopaminergic neuron embryo stage 2 gene function genetic regulatory protein improved insight interest knock-down loss of function nervous system development neurogenesis notch protein relating to nervous system research study response retinal rods

项目摘要

The major questions we focus on are: 1) What are the upstream regulatory components that specify the animal pole domain (APD) of the sea urchin embryo, which contains nerves and cells bearing long, immotile cilia? 2) What are the signals transmitted during early cleavage and blastula stages that are required for endomesoderm specification and timely gastrulation? 3) What is the molecular basis for the embryos ability to change its morphology to adapt to changing food concentrations? In each case, we assay the expression of all predicted genes in the genome in order to gain a complete understanding of the gene regulatory networks that underlie these processes. Six3 is necessary and sufficient for APD specification (45%) (Zheng Wei, Junko Yaguchi, Shunsuke Yaguchi, Lynne Angerer) During the last several years, using bioinformatics approaches and molecular screens, we identified many genes encoding regulatory proteins expressed specifically in the primary neurogenic domain of the sea urchin embryo. Two of the earliest, FoxQ2 and Six3, control early decisions in ectodermal patterning. Experiments conducted during 2008 showed that Six3 is necessary and sufficient for all known features of APD development, including all neurons formed during embryogenesis and for the large majority of APD-specific regulatory proteins previously identified, including FoxQ2. We identified the full regulatory repertoire of genes that depends on Six3, and discovered that Six3 can also suppress canonical Wnt and TGF-beta signals that pattern the rest of the embryo. Our work revealed that Six3 function in the sea urchin embryo APD and the vertebrate forebrain share some features. Because our studies have identified many additional regulatory genes that are Six3-dependent and expressed in the APD, they are likely to facilitate understanding of vertebrate forebrain development. A manuscript reporting this work (Wei et al.) is in review. FoxQ2 is a checkpoint coordinating cell fate specification along the primary and secondary developmental axes (5%)(Shunsuke and Junko Yaguchi) FoxQ2 negatively regulates TGF-beta signaling, which is required for patterning along the secondary axis of all regions of ectoderm except that at the animal pole. In order for TGf-beta signaling to occur, FoxQ2 must be eliminated from the lateral ectoderm, an event that depends on canonical Wnt, the primary axis signaling system. FoxQ2 provides a checkpoint coordinating patterning along the primary and secondary axes (Yaguchi et al., Developmental Cell 14, 97-107, 2008). It also functions in the differentiation of different cell types in the APD and may mediate Six3s role. Whole-genome screens show that is required for the expression of nearly 500 genes. The achievements this year provide strong impetus for determining the structure and cell type specificity of the FoxQ2-dependent GRN, as well as the Six3-dependent GRN, in the neurogenic domain at the animal pole. ActivinB/ALK4/5/7 and Delta/Notch are early signals that induce specification of endomesoderm and endoderm (25%)(Adi Sethi, Radhika Wikramanayake) Previous experiments showed that both ectopic and normal endomesoderm induction requires ActivinB and that this signaling factor executes all of the endomesoderm inducing functions of the previously unknown, but long-sought, early micromere signal. His work is the first to connect ActivinB signaling to specific components of an endomesoderm gene regulatory (GRN) network, the largest and best developed in any embryo, and it has led to new insights into its operation. During this year additional experiments carried out in response to reviews have confirmed previous work and an improved, revised manuscript is in review. Previous work blocking Notch function reveal new functions for this pathway in separating two primary germ layers, the endoderm and mesoderm, within the endomesodermal field. During 2008, we found that micromere Delta signals not only aactivate gcm, the cardinal pigment cell GRN regulator, but also to repress several critical genes required for early endoderm specification. Delta signaling to endoderm is earlier than previously reported and necessary for expression of early endoderm specification genes. Because macromere Delta depends on prior micromere Delta signals, new methods to knock down it down in specific blastomeres have been developed. These include injection of a morpholino blocking Delta translation in single blastomeres or inducing loss of Delta transcription in specific blastomeres using antisense intron RNAs. Preliminary experiments suggest that both methods will be useful. Dopaminergic neurons regulate the embryos response to food density (25%) (Diane Adams) A new line of investigation aims to understand the molecular pathways by which larvae sense and respond to the concentration of food by adjusting arm structure to maximize food intake. It is a high-risk project that bridges larval developmental biology, larval physiology and the ecology of larval dispersal. Diane has found that changes in the post-oral skeletal rod length in response to food density is regulated by dopamine and that dopaminergic neurons are positioned correctly at the base of the post-oral arms to regulate the sensory response. To determine the components of the food sensing pathway, she is assaying the whole geneme reponse to food concentration and to dopamine antagonists and agonists using microarrays. She has also established a collaboration with Dr. Andrew Cameron at Caltech to search for G-coupled protein receptors that might function in larval sensing of food during larval stages.

我们关注的主要问题是：1）指定海胆胚胎的动物极域（APD）的上游调节组件是什么，其中包含长长，不含纤毛的神经和细胞？ 2）内胚层规范所需的裂解和囊肿阶段期间传播的信号是什么？ 3）胚胎改变其形态以适应不断变化的食物浓度的能力的分子基础是什么？在每种情况下，我们分析了基因组中所有预测基因的表达，以便对这些过程的基因调节网络有完整的了解。 SIX3是APD规格的必要条件（45％）（Zheng Wei，Junko Yaguchi，Shunsuke Yaguchi，Lynne Angerer）在过去的几年中，使用生物信息学方法和分子筛选，我们确定了许多编码调节蛋白的基因，这些基因专门在海胆胚胎的原发性神经发生结构域中表达。最早的两个FOXQ2和SIX3最早控制了外胚层图案的早期决定。 2008年期间进行的实验表明，Six3对于APD发育的所有已知特征，包括在胚胎发生过程中形成的所有神经元以及以前鉴定出的大多数APD特异性调节蛋白（包括FOXQ2）的大多数神经元。我们确定了取决于六3的基因的完整调节曲目，并发现Six3还可以抑制规范的Wnt和TGF-beta信号，这些信号对胚胎的其余部分进行了模式。我们的工作表明，海胆胚胎APD的六个功能和脊椎动物前脑具有某些特征。由于我们的研究已经确定了六个依赖性并在APD中表达的许多其他调节基因，因此它们很可能有助于理解脊椎动物前脑发育。报告这项工作的手稿（Wei等人）正在审查中。 FOXQ2是沿主要和次要发育轴（5％）的检查点协调细胞命运规范（Shunsuke和Junko Yaguchi） FOXQ2负调控TGF-β信号传导，这是沿着外胚层所有区域的二级轴构图所必需的，除了在动物极处。为了使TGF-beta信号发生，必须从外侧外胚层中消除FOXQ2，这一事件取决于主要轴信号系统的规范Wnt。 FOXQ2提供了沿主要轴和次级轴的检查点协调模式的（Yaguchi等人，发育单元格14，97-107，2008）。它还在APD中不同细胞类型的分化中起作用，并可能介导SIX3S角色。全基因组筛选显示了近500个基因表达所需的。今年的成就为确定动物极点神经源性结构域中FOXQ2依赖性GRN的结构和细胞类型特异性提供了强大的动力。 activinb/Alk4/5/7和Delta/Notch是诱导内胚层和内胚层规范的早期信号（25％）（Adi Sethi，Radhika Wikramanayake）先前的实验表明，异位和正常内胚层诱导都需要激活素，并且该信号因子执行了先前未知但长期久的早期微米信号的所有内胚层诱导功能。他的工作是第一个将Activinb信号传导与内胚层基因调节（GRN）网络的特定组件连接起来的工作，该网络是任何胚胎中最大，最佳开发的，这导致了对其操作的新见解。在这一年中，针对审查进行了其他实验，证实了先前的工作，并进行了改进的修订手稿。先前的工作阻止Notch函数在内胚层场中揭示了该途径的新功能，即在内胚层和中胚层分离两个主要的细菌层。在2008年期间，我们发现Micromere Delta信号不仅是Airtivate GCM（基本色素细胞GRN GRN调节剂），而且还抑制早期内胚层规范所需的几个关键基因。向内胚层的增量信号传导比先前报道的早期报道要早，这对于表达早期内胚层规范基因所必需。由于Macromere Delta取决于先前的Micromere Delta信号，因此已经开发了在特定的胚泡中将其击倒的新方法。这些包括使用反义含有RNA在特定胚胎中注入单胞菌中的形态封闭的三角洲翻译或在特定胚胎中诱导三角洲转录的损失。初步实验表明两种方法都将有用。多巴胺能神经元调节胚胎对食物密度的反应（25％）（黛安·亚当斯）一项新的调查旨在了解幼虫通过调节手臂结构以最大化食物摄入量来对食物的浓度的分子途径来理解分子途径。这是一个高风险的项目，它桥接了幼虫发育生物学，幼体生理学和幼虫分散的生态。黛安（Diane）发现，响应食物密度的术后骨骼杆长度的变化受多巴胺调节，多巴胺能神经元正确定位在后臂的底部，以调节感觉反应。为了确定食物传感途径的组成部分，她正在使用微阵列分析整个基因对食物浓度和多巴胺拮抗剂和激动剂的反应。她还与加州理工学院（Caltech）的安德鲁·卡梅伦（Andrew Cameron）博士建立了合作，以寻找可能在幼虫阶段在食物的幼虫感应中起作用的G耦合蛋白受体。