Decoding lineage and fate specification in the C. elegans embryo

摘要 由发育调节的转录因子进行的组合调节在 定义哪些基因在发育和疾病期间在每个细胞中受到调控，并允许相同的因素 在不同的细胞中扮演不同的角色。线虫胚胎是全面研究线虫的理想系统 血统历史在细胞命运的上下文依赖调控中的作用，因为它具有不变的血统和 强大的实验工具。我们最近开发了自动谱系跟踪和表达式映射 线虫胚胎发生的方法，并建立了“线虫表达模式”(EPIC) 包含超过250个转录因子(TF)表达的荧光记者的数据库 在发育中的胚胎的每一个细胞中。在我们过去的工作中，我们已经展示了这种资源作为一种 定义控制发展的机制的起点。同时我们将继续映射该表达式 对于新的监管机构，这项建议的主要重点是利用这个数据库和我们的方法来应对糟糕的 理解发育生物学中的问题。1)细胞如何知道它们的谱系历史和翻译 将这些信息输入到正确的终端细胞命运中？我们已经确定了一组大约15个特定于血统的TF，他们 表达区分不同组织的不同组织的祖细胞群 卵裂球。我们计划测试这些函数是否单独和组合在一起是必要和充分的 确定这些祖细胞的谱系特征，并通过成像和基因组学确定它们的靶点 功能缺失和功能获得突变体的分析。我们将通过对CI的详细分析来补充这一点- 控制末端分化基因的调控序列，以识别其上游调控基因。这些 “自上而下”和“自下而上”的方法最终应该汇聚成一个共同的监管网络。2)什么 机制允许同一调节器(S)在不同发育阶段用于不同目的 背景，比如不同的血统？我们已经确定了两组基因，分别在后部或后部表达 细胞分裂后的前子细胞，两者都受Wnt途径的调控。我们将联合起来 增强子精细作图，合成增强子的表达作图，以及全基因组结合分析 WNT调节的Tf POP-1/TCF，以确定每个基因对此的反应(激活或抑制) 途径在不同的细胞中受到不同的调控。3)基因和增强子的冗余是如何 影响发育健壮性？冗余在早期胚胎中非常常见，我们将 检验这种冗余存在的假设，以确保在面对环境的情况下强劲发展 通过对不同条件下的突变体进行详细的表型分析来确定变异性。总而言之，我们提出的工作将 开始完成早期胚胎调控网络，并回答有关原则的重要问题 很可能在动物中保存下来的发育。

ABSTRACT Combinatorial regulation by developmentally regulated transcription factors play a central role in defining which genes are regulated in each cell during development and disease, and allows the same factors to play different roles in different cells. The C. elegans embryo is an ideal system for a comprehensive study of the role of lineage history in the context-dependent regulation of cell fate because of its invariant lineage and powerful experimental tools. We recently developed automated lineage tracing and expression mapping methods for C. elegans embryogenesis and have built the “Expression Patterns in Caenorhabditis” (EPIC) database that contains the expression of over 250 fluorescent reporters for transcription factor (TF) expression in every cell of developing embryos. In our past work, we have shown the potential for this resource as a starting point for defining mechanisms controlling development. While we will continue to map the expression of novel regulators, the main focus of this proposal is to use this database and our methods to address poorly understood questions in developmental biology. 1) How do cells know their lineage history and translate this information into correct terminal cell fates? We have identified a set of ~15 lineage-specific TFs whose expression distinguishes a group of progenitor cells for diverse tissues descended from the “ABpxp” blastomeres. We plan to test whether these TFs are necessary and sufficient individually and in combination to specify the lineage identity of these progenitor cells, and to identify their targets, by imaging and genomics analysis of loss and gain-of-function mutants. We will complement this by a detailed analysis of the cis- regulatory sequences controlling terminal differentiation genes to identify their upstream regulators. These “top-down” and “bottom-up” approaches should eventually converge to a common regulatory network. 2) What mechanisms allow reuse of the same regulator(s) for different purposes in different developmental contexts, such as different lineages? We have identified two sets of genes, expressed in either posterior or anterior daughter cells after cell division, both of which are regulated by the Wnt pathway. We will combine enhancer fine-mapping, expression mapping of synthetic enhancers, and genome-wide binding assays for the Wnt-regulated TF POP-1/TCF to determine how each gene's response (activation or repression) to this pathway is regulated differently in different cells. 3) How does redundancy of genes and enhancers influence developmental robustness? Redundancy is extremely common in the early embryo, and we will test the hypothesis that this redundancy exists to ensure robust development in the face of environmental variability by detailed phenotyping of mutants in different conditions. In summary, the work we propose will begin to complete the early embryonic regulatory network and answer important questions about principles of development that are likely to be conserved across animals.