Collaborative Research: Enhancers and the Convergent Evolution of Limb Deduction in Squamates

合作研究：有鳞动物肢体扣除的增强子和趋同进化

• 批准号: 1754417
• 负责人: Brant Faircloth
• 金额: $ 21.22万
• 依托单位: Louisiana State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1754417&HistoricalAwards=false
• 关键词: Collaborative; Research; Enhancers; Convergent; Evolution

Although the development of arms and legs is a key evolutionary innovation that allowed animals to move onto land, many species of snakes and lizards subsequently lost these limbs. Indeed, limb loss has evolved at least 24 times among lizards. How limb loss has occurred repeatedly in lizard lineages is a question that can be addressed with the replicated natural experiments that will be the focus of this study. This project will reveal whether the same or different genetic changes are associated with limb loss among these different lizard groups. This research will combine technologies from different fields of biology in novel ways to explore this extraordinary example of repeated evolution and gain new insight into genome function. This project will greatly improve knowledge of the genes and genetic control elements involved in limb development. Additionally, in the course of this project, thousands of genetic regions that are shared by humans and other animals will be characterized. Therefore, project data will deliver insights that can be applied to better understand other functional elements within the genomes of humans and different animals. The methods developed will pave the way for other researchers who wish to leverage the power of natural evolutionary experiments to understand genome function. Finally, materials for educators will be developed for use in teaching students how information is encoded in genomes and how natural evolutionary experiments can be used to reveal how genetic sequences yield unique biological diversity.This project will use chromatin-immunoprecipitation followed by high-throughput DNA sequencing (ChIP-seq) to identify regulatory elements (enhancers) in squamate genomes that control the embryonic development of the limbs. This information will be used to design squamate-specific DNA sequence capture baits targeting these regulatory regions. Additional capture baits will be designed to target the coding regions of genes active in the developing limbs. Together, these baits will be used to survey both noncoding regulatory elements and coding exons of key developmental genes from every lineage of limb-reduced squamates. DNA sequence changes in limb enhancers and limb development genes will be compared in limb-reduced species and their fully limbed relatives, and functional changes in regulatory activity will be assessed in vivo with transgenic mouse reporter assays. This will provide insight into the relative frequency of regulatory changes compared to gene coding sequence changes contributing to the evolution of limb loss. Finally, the functional evolution of an ancient amniote limb-genital enhancer for the gene Tbx4 will be dissected molecularly by taking advantage of sequence variation across the multiple lineages of limb-reduced species. This will grant insight into the fate of enhancers that have lost their regulatory roles and determine whether these elements are recruited to new functions.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.

虽然手臂和腿的发展是一个关键的进化创新，使动物能够在陆地上移动，但许多种类的蛇和蜥蜴随后失去了这些肢体。事实上，在蜥蜴中，断肢至少进化了24次。如何在蜥蜴谱系中反复发生肢体缺失是一个可以通过复制自然实验来解决的问题，这将是本研究的重点。该项目将揭示是否相同或不同的基因变化与这些不同蜥蜴群体的肢体缺失有关。这项研究将联合收割机以新颖的方式结合来自不同生物学领域的技术，探索这个重复进化的非凡例子，并获得对基因组功能的新见解。该项目将极大地提高基因和基因控制元素参与肢体发育的知识。此外，在这个项目的过程中，人类和其他动物共享的数千个基因区域将被表征。因此，项目数据将提供可用于更好地了解人类和不同动物基因组中其他功能元件的见解。开发的方法将为其他希望利用自然进化实验的力量来了解基因组功能的研究人员铺平道路。最后，将为教育工作者编写教材，用于教授学生如何在基因组中编码信息，以及如何利用自然进化实验来揭示基因序列如何产生独特的生物多样性。该项目将使用染色质免疫沉淀和高通量DNA测序（ChIP-seq）来确定控制肢体胚胎发育的鳞片基因组中的调节元件（增强子）。这些信息将用于设计靶向这些调控区域的鳞片特异性DNA序列捕获诱饵。额外的捕获诱饵将被设计成靶向发育中的肢体中活跃的基因编码区。总之，这些诱饵将被用来调查非编码调控元件和编码外显子的关键发育基因的每一个血统的肢体减少有鳞动物。将在肢体减少的物种及其完全肢体的亲属中比较肢体增强子和肢体发育基因的DNA序列变化，并将使用转基因小鼠报告基因测定在体内评估调节活性的功能变化。这将提供洞察的相对频率的监管变化相比，基因编码序列的变化，有助于肢体丧失的演变。最后，一个古老的古龙的肢体生殖器增强子的基因Tbx 4的功能进化将解剖分子利用跨多个谱系的肢体减少物种的序列变异。该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。