Analysis of a Model Developmental Gene Regulatory Network

在发育过程中，多细胞生物是由一个单细胞——受精卵产生的。反复的细胞分裂产生了由许多细胞组成的胚胎。这些细胞逐渐承担不同的功能，并自行组织成组织和器官。理解发育具有重要的科学意义，因为这是新一代多细胞生物产生的过程。值得注意的是，胚胎发育的整个蓝图包含在基因组的DNA序列中。现代生物学的一个核心挑战是解释基因组信息如何被“解码”以控制发育过程。这里描述的研究项目解决了这个问题，使用海胆胚胎作为一个方便的实验模型。它通过使用新的方法来检查基因组DNA的物理结构，监测胚胎中各种基因的工作时间和地点，并确定这些基因之间的相互作用，来检查这些海洋生物骨骼形成的编码基因。这项工作将阐明在单个细胞发育成这些生物体的关键部分的过程中，基因组中包含的信息被破译的机制。发育生物学的核心挑战是将基因与解剖学联系起来。驱动发育的差异基因表达程序在很大程度上受调控基因（即编码转录因子的基因，或TFs）和TFs结合的非编码DNA序列的复杂网络控制。这些基因调控网络（grn）是分析遗传控制和发育进化的有力工具。目前，我们对grn与直接形成胚胎组织和器官（形态发生）的过程之间的联系了解非常有限。这里描述的项目利用海胆胚胎解决了这个问题，这是一个卓越的GRN生物学和形态发生分析系统。该项目利用了任何发育生物体中最完整的GRN之一，GRN驱动初级间充质细胞（pmc）的一系列细胞行为，并最终形成胚胎骨架。该研究计划采用“自上而下”的方法来剖析由Alx1控制的基本子电路的结构，Alx1是骨骼生成GRN中的关键TF。这些研究利用基因敲低、mRNA过表达、基因表达谱和基于生物信息学的方法。与此同时，一种“自下而上”的方法被采用，该方法使用报告基因分析来剖析控制形态发生效应基因转录的非编码调控DNA序列。该信息用于引导进入上游网络电路。最后，该项目扩展了基因敲低筛选，以鉴定和表征从pmc富集转录物和Alx1靶点的全基因组分析中出现的新的形态发生效应物。

Non-Technical ParagraphIn development, multicellular organisms arise from a single cell - a fertilized egg. Repeated cell divisions create an embryo composed of many cells. These cells gradually take on distinct functions and organize themselves into tissues and organs. Understanding development is of cardinal scientific importance, as this is the process by which new generations of multicellular organisms arise. Remarkably, the entire blueprint for embryonic development is contained in the DNA sequence of the genome. A central challenge of modern biology is to explain how this genome information is "decoded" to control the process of development. The research project described here addresses this question, using the sea urchin embryo as a convenient experimental model. It examines the genes that encode the formation of the skeleton of these sea creatures through use of new methods to examine the physical structure of genomic DNA, to monitor when and where in the embryo various genes work, and to determine interactions among these genes. The work will shed light on the mechanisms by which the information contained in the genome is deciphered during the development of a single cell into a critical part of these organisms. Technical ParagraphThe central challenge of developmental biology is to link genes to anatomy. The programs of differential gene expression that drive development are largely controlled by complex networks of regulatory genes (i.e., genes that encode transcription factors, or TFs) and the non-coding DNA sequences to which TFs bind. Such gene regulatory networks (GRNs) are powerful tools for analyzing the genetic control and evolution of development. At present, we have a very limited understanding of the connections between GRNs and the processes that directly shape embryonic tissues and organs (morphogenesis). The project described here addresses this question using the sea urchin embryo, a pre-eminent system for both GRN biology and the analysis of morphogenesis. The project leverages one of the most complete GRNs in any developing organism, a GRN that drives a suite of cell behaviors by primary mesenchyme cells (PMCs) and culminates in the formation of the embryonic skeleton. The research plan takes a "top-down" approach to dissect the architecture of an essential subcircuit controlled by Alx1, a key TF in the skeletogenic GRN. These studies utilize gene knockdowns, mRNA overexpression, gene expression profiling, and bioinformatics-based approaches. In parallel, a "bottom-up" approach is taken that uses reporter gene assays to dissect non-coding regulatory DNA sequences that control the transcription of morphogenetic effector genes. This information is used to bootstrap into the upstream network circuitry. Lastly, the project extends a gene knockdown screen to identify and characterize new morphogenetic effectors that have emerged from genome-wide profiling of PMC-enriched transcripts and Alx1 targets.