DISSERTATION RESEARCH: Mutant models reveal latent developmental potential with roles in evolutionary change

这个项目将调查导致鱼鳍上精细的骨骼图案的遗传因素。骨骼模式的细化是骨骼数量和连接增加的结果，这种研究对于理解进化和发育过程都很重要。在进化过程中，骨头端到端排列数量的增加有助于将鳍转化为四肢，并有助于灭绝脊椎动物对陆地的殖民。在发育过程中，正常的功能和运动依赖于适当的骨骼数量和连接的形成，而发育缺陷可能会导致严重的损害。目前，导致鳍骨架形成肢体的机制尚不清楚。为了确定涉及的遗传因素，研究人员将研究模式物种斑马鱼的品系，这些品系具有导致新鳍骨形成的基因变化。他们还将研究从肩部延伸出不同数量的骨骼的两个物种--加和弓鳍的自然发生的变异。这些互补的方法将揭示调控鳍和肢体骨骼生长和形成的机制，并展示简单的基因变化如何导致鳍形成肢体。了解骨骼模式的形成和发育机制对人类健康和发育异常具有重要意义，并可能为陆地生命的进化提供线索。此外，该项目将为美洲原住民本科生提供研究机会。虽然四足脊椎动物增加了肢体近端-远端(P-D)轴上的骨骼数量，但硬骨鱼(如斑马鱼)减少了胸鳍骨架；沿P-D轴只发现了一根长骨。研究人员对斑马鱼进行了正向突变筛选，并分离出一种新的突变体，它通过一个类似于四足动物四肢发育过程的过程，沿着鳍的P-D轴形成额外的骨骼。这项拟议的研究将把突变分析的结果扩展到自然种群，以确定突变是否通过激活祖先的遗传机制来产生多余的骨骼。通过RNA测序的转录组图谱将被用来比较野生型和突变型斑马鱼以及在P-D轴精加工方面表现出自然变异的两个物种胸鳍发育中的基因表达水平。这一分析将揭示与P-D轴细化相关的差异基因表达模式。将在斑马鱼身上测试差异表达的基因，以确定它们的修饰表达是否对鳍的形态有影响。这项研究将阐明加性表型变化是如何在发育过程中出现的，并确定在进化和疾病中形成额外的鳍/肢骨的候选途径。

This project will investigate the genetic factors that cause elaborated bone pattern in fish fins. Bone pattern elaboration results from increases in bone numbers and connections, and such research is important for understanding processes of both evolution and development. In evolution, increase in the number of bones arranged end-to-end helped transform fins into limbs and aided the colonization of land by extinct vertebrates. In development, normal function and movement depends upon formation of the proper bone numbers and connections, and defective development can lead to severe impairment. Currently, the mechanisms that led to elaboration of the fin skeleton to form limbs are unknown. To identify the genetic factors involved, the researchers will study strains of the model species, zebrafish, that have genetic changes that result in the formation of new fin bones. They will also investigate naturally occurring variation in two species, gar and bowfin, that have different numbers of bones extending from the shoulder. These complementary approaches will reveal the mechanisms regulating growth and formation of the skeleton of fins and limbs, and show how simple genetic changes can lead to the formation of limbs from fins. Understanding the mechanisms of bone pattern elaboration and development has implications for human health and developmental abnormalities, and may provide clues for the evolution of life on land. Additionally, the project will provide research opportunities for Native American undergraduates. While tetrapod vertebrates have increased the number of bones along the proximo-distal (P-D) axis of the limb, teleost fishes such as the zebrafish have reduced the pectoral fin skeleton; only a single long bone is found along the P-D axis. The researchers performed a forward mutagenesis screen in zebrafish and isolated a novel mutant that forms supernumerary bones along the P-D axis of the fin through a process similar to that seen in the development of tetrapod limbs. The proposed research will extend results from mutant analysis to natural populations to determine if the mutation is producing supernumerary bones by activating ancestral genetic mechanisms. Transcriptome profiling by RNA sequencing will be used to compare gene-expression levels in the developing pectoral fins of wild-type and mutant zebrafish, as well as two species that exhibit natural variation in P-D axis elaboration. This analysis will reveal patterns of differential gene expression that correlate with P-D axis elaboration. Differentially expressed genes will be tested in the zebrafish to determine if their modified expression has an effect on fin morphology. This study will elucidate how additive phenotypic changes arise in development and identify candidate pathways by which additional fin/limb bones are formed in both evolution and disease.