Gene expression divergence during Drosophila head development on single cell resolution

单细胞分辨率下果蝇头部发育过程中的基因表达差异

• 批准号: 417980402
• 负责人: Dr. Nico Posnien
• 金额: --
• 依托单位: Abteilung Entwicklungsbiologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2019
• 资助国家: 德国
• 起止时间: 2018-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/417980402?language=en
• 关键词: Gene; expression; divergence; during; Drosophila

Understanding the processes involved in the origin of the diversity of life forms observed on earth is a major aim of biological research. Adult morphology is defined during embryonic and postembryonic development by the action of developmental gene products, which are organized in gene regulatory networks (GRNs). Natural variation in the expression and thus the regulation of developmental genes has been shown to be a main driver for morphological diversification. While the developmental underpinnings of differences in relatively simple traits are being revealed, a comprehensive understanding of the molecular changes responsible for morphological divergence of complex traits is missing to date. To close this gap, we study the evolution of head development in three closely related Drosophila species. The insect head is a complex structure that consists of the main sensory organs (e.g. the compound eye and antennae) to perceive environmental information and the brain to process information to trigger behavioral responses. The Drosophila head develops from a composite eye-antennal imaginal disc that undergoes major patterning, growth and differentiation processes during larval stages. We and others have previously quantified differences in adult head size and shape among the three Drosophila species D. melanogaster, D. simulans and D. mauritiana. To identify candidate genes involved in the observed variation, we generated a comprehensive comparative transcriptomic dataset by sequencing entire eye-antennal imaginal discs throughout development (i.e. bulk-RNAseq). Based on this data we revealed a high level of context dependent gene expression and regulation recapitulating the composite nature of the developing eye-antennal imaginal discs. The identification of clear candidate genes and developmental processes, however, is hampered because the bulk-RNAseq data does not provide sufficient tissue specific resolution. Therefore, we propose here to study gene expression dynamics and divergence on single cell level using single cell mRNA sequencing (scRNAseq).We will establish a single cell expression atlas for five stages of eye-antennal imaginal disc development for D. melanogaster, D. simulans and D. mauritiana. This data will be used to reveal key developmental genes and regulatory interactions in specific cell clusters representing different regions of the developing head. The interspecific comparison of observed cell clusters with respect to gene expression and GRN architecture will allow identifying diverging cell populations. Two of these cell clusters will be functionally validated applying CRISPR/Cas9 mediated genome editing and re-sequencing of cluster specific cells. With the proposed project we will gain major insights into the development and evolution of the Drosophila head on an unprecedented resolution. Additionally, we will contribute to a broader mechanistic understanding of context dependent gene expression and regulation.

了解地球上观察到的生命形式多样性起源的过程是生物学研究的一个主要目标。成体形态在胚胎和胚胎后发育过程中由发育基因产物的作用决定，这些基因产物在基因调控网络（grn）中组织。表达的自然变异以及发育基因的调控已被证明是形态多样化的主要驱动因素。虽然相对简单性状差异的发育基础正在被揭示，但迄今为止，对复杂性状形态差异的分子变化仍缺乏全面的理解。为了缩小这一差距，我们研究了三个密切相关的果蝇物种的头部发育进化。昆虫的头部是一个复杂的结构，包括主要的感觉器官（如复眼和触角）来感知环境信息，以及大脑来处理信息以触发行为反应。果蝇的头部由一个复合的眼-触角影像盘发育而成，在幼虫阶段经历了主要的模式、生长和分化过程。我们和其他人之前已经量化了三种果蝇（D. melanogaster， D. simulans和D. mauritiana）在成年头部大小和形状上的差异。为了确定与观察到的变异相关的候选基因，我们通过对整个发育过程中的眼天线成像盘（即bulk-RNAseq）进行测序，生成了一个全面的比较转录组数据集。基于这些数据，我们揭示了高水平的环境依赖基因表达和调控，概括了发育中的眼天线成像盘的复合性质。然而，明确候选基因和发育过程的鉴定受到阻碍，因为大量rnaseq数据不能提供足够的组织特异性分辨率。因此，我们建议使用单细胞mRNA测序（scRNAseq）来研究基因在单细胞水平上的表达动态和分化。我们将建立眼触角影像盘发育五个阶段的单细胞表达图谱，包括黑腹龙、拟象龙和毛里求斯龙。这些数据将用于揭示关键的发育基因和代表发育头部不同区域的特定细胞簇中的调节相互作用。与基因表达和GRN结构相关的观察到的细胞簇的种间比较将允许识别不同的细胞群。其中两个细胞簇将通过CRISPR/Cas9介导的基因组编辑和簇特异性细胞的重新测序进行功能验证。通过这个项目，我们将以前所未有的分辨率深入了解果蝇头部的发展和进化。此外，我们将有助于对环境依赖基因表达和调控的更广泛的机制理解。