Evolution of segmentation in chordates: dissecting the genetic mechanism of somitogenesis in the basal chordate, amphioxus

非技术性的。了解组成有机体基因组的数千个基因(基因型)如何相互作用，创造出共同组成有机体(表型)的组织和器官，是美国国家研究委员会确定的五个“21世纪生物学的重大挑战”之一。剖析这种基因-表型关系具有重要的现实意义，也是大量研究工作的重点。重要的是，它是了解生物体如何进化的关键，并对健康科学、农业、生物技术和生态学具有广泛的影响。为了解决基因和表型的关系，这项研究使用了文昌鱼，这是一种简单的类似鱼的动物，其祖先在5亿年前从导致脊椎动物的进化路线中分离出来。具体的问题是，文昌鱼基因组是如何产生分段的身体肌肉的(类似于脊椎动物(如鱼)两侧重复的肌肉，允许左右移动)。技术上的。这项研究将阐明介导肌肉分割的基因网络，并在单细胞水平上展示该网络如何控制细胞分割成离散的片段(体节)。与脊椎动物的比较将表明，随着脊椎动物变得更大和更复杂，这种遗传机制发生了怎样的变化。文昌鱼的肌节延伸到整个身体，但在脊椎动物中，头部没有肌节。脊椎动物是失去了头部节段，还是文昌鱼的新奇事物，这是有争议的。在文昌鱼和脊椎动物之间，肌肉分割的大部分基因网络似乎是保守的--在两者中，“时钟”规定了肌肉分割的形成时间，而Notch/Delta信号介导了肌肉分割。然而，在脊椎动物中，分泌蛋白质的梯度，主要是FGFs，指定了节段边界的形成位置，但在文昌鱼中，只有头部节段，而不是躯干或尾巴中的那些，需要FGFs。为了验证随着脊椎动物变大而失去头部节段，成纤维细胞生长因子梯度从头部到后节的理论，文昌鱼发育过程中的关键信号通路(Wnt/β-catenin；成纤维细胞生长因子，Notch)将在特定的发育时间被化学抑制剂和激活剂干扰。基因敲除和过度表达将显示基因层级。对分割形态的影响将通过连续的块面扫描电子显微镜(SBSEM)在三维中重建，如果每隔一段时间进行，它可以在单细胞水平上显示形态发生的步骤。研究人员将让本科生和高中生直接参与该项目，还将与斯克里普斯海洋研究所的桦树水族馆和圣地亚哥县教育办公室合作，为高中教师主持研讨会。这些研讨会将使用当地可获得的海洋无脊椎动物向教师展示如何指导学生进行实验设计和数据分析-这是培养下一代科学家的基本技能。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Non-technical. Understanding how the thousands of genes that make up an organism’s genome (the genotype) interact to create the tissues and organs that together make up that organism (the phenotype) is one of the five ‘grand challenges of 21st century biology’ identified by the National Research Council. Dissecting this genotype-phenotype relationship is of great practical importance and is the focus of enormous research efforts. Importantly, it is key to understanding how organisms evolve and has wide-ranging implications for the health sciences, agriculture, biotechnology, and ecology. To address the genotype-phenotype relationship, this research uses amphioxus, a simple fish-like animal whose ancestors separated from the evolutionary line leading to vertebrates half a billion years ago. The specific question asked is how the amphioxus genome produces the segmented body muscles (similar to the repeating muscles along each side of a vertebrate such as a fish that allow side to side movement). Technical. The research will elucidate the gene network mediating muscle segmentation, and show at the single-cell level how this network controls the partitioning of cells into discrete segments (somites). Comparisons with vertebrates will show how this genetic mechanism has changed as vertebrates became larger and more complex. In amphioxus, muscle segments extend the full length of the body, but in vertebrates, there are none in the head. It is controversial whether vertebrates lost head segments or they are an amphioxus novelty. Much of the gene network for muscle segmentation appears to be conserved between amphioxus and vertebrates—in both, a “clock” specifies when segments will form, and Notch/Delta signaling mediates segmentation. However, in vertebrates, gradients of secreted proteins, chiefly Fgfs, specify where segment boundaries will form, but in amphioxus only the head segments, not those in the trunk or tail, require Fgfs. To test the theory that as vertebrates grew larger and lost head segments, the Fgf gradient was co-opted from the head to the posterior segments, key signaling pathways (Wnt/β-catenin; Fgf, Notch) in developing amphioxus will be perturbed with chemical inhibitors and activators at specific developmental times. Gene knockdown and overexpression will show gene hierarchy. Effects on morphology of segmentation will be reconstructed in 3-D by serial blockface scanning electron microscopy (SBSEM), which when done at intervals can show the steps in morphogenesis at the single cell level. Researchers will involve undergraduates and high school students directly in the project and also partner with the Birch Aquarium at Scripps Institution of Oceanography and the San Diego County Office of Education to lead workshops for high school teachers. These workshops will use locally-available marine invertebrates to show teachers how to instruct students in experimental design and data analysis—skills that are fundamental to training the next generation of scientists.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.