Defining the genetic and cellular basis of morphological diversity in Drosophila

项目摘要 这项提案的总体目标是确定改变复杂三维结构的遗传变异。 并确定这些基因如何对细胞数量、形状和行为产生影响 在发展过程中。在形态发生中起重要作用的发育基因以复杂的方式联系在一起 层次结构，但我们目前还不知道如何在基因调控的背景下优先考虑变异， 网络因此，我将在基因调控网络的水平上进行进化案例研究， 确定有助于物种之间形态多样性的基因类型， 对单个细胞的影响。为了实现这个目标，我需要一个系统， 一个数量性状的基础是很好理解的，关键的变异可以在分辨率映射 个体基因黑腹果蝇分支的后叶特别适合于 将遗传变异与形态发生联系起来。最近的工作已经确定了许多组成部分， 控制后叶发育的基因调控网络，包括模式化分子（转录 因子和信号分子）和细胞效应物（细胞骨架调节剂）。此外，混合动力车 这三个物种都是可行的，并表现出中间叶形态，这使得严格的映射， 基因组区域导致了它们截然不同的表型。在目标1中，我将映射序列 负责后叶形态的定量变化。我将在物种中使用RNA-seq， 杂交以鉴定含有表达变异的基因，期望这些基因中的一些 有助于后叶的形态多样性。然后，我将使用 基于CRISPR的互补测试。在目标2中，我将确定如何图案化分子和细胞 效应物影响单个细胞的数量、形状和行为。我会用细胞标记和成像 测量来自突变体、不同物种和 杂交种通过完成这一建议，我将连接基因调控网络的变化， 决定细胞大小和形状的分子机制，这将发展后叶作为一个总理， 系统的形态发展和进化的系统级研究。

PROJECT SUMMARY The overall goal of this proposal is to determine the genetic variants that alter complex three-dimensional morphologies and to determine how these genes exert their influence on cell number, shape and behavior during development. Developmental genes that are important in morphogenesis are linked in complex hierarchies, yet we do not currently know how to prioritize variants within the context of a gene regulatory network. I will therefore undertake an evolutionary case study at the level of a gene regulatory network to determine the types of genes that contribute to morphological diversification between species and characterize their effects on individual cells. To accomplish this goal, I require a system where the gene regulatory network underlying a quantitative trait is well-understood, and where critical variants can be mapped at the resolution of individual genes. The posterior lobe in the Drosophila melanogaster clade is particularly well-suited to connecting genetic variation to morphogenetic outcomes. Recent work has identified many components of the gene regulatory network controlling posterior lobe development, including patterning molecules (transcription factors and signaling molecules) and cellular effectors (cytoskeletal regulators). Furthermore, hybrids between all three species are viable and exhibit intermediate lobe morphologies, which allows for rigorous mapping of genomic regions contributing to their drastically different phenotypes. In Aim 1, I will map sequences responsible for quantitative variation in posterior lobe morphology. I will use RNA-seq in species and hybrids to identify genes containing expression variation, with the expectation that some of these genes contribute to morphological diversity in the posterior lobe. I will then validate strong candidates using a CRISPR-based complementation test. In Aim 2, I will determine how patterning molecules and cellular effectors influence the number, shape and behavior of individual cells. I will use cell labeling and imaging to measure cell number, size and behavior in posterior lobes derived from mutants, different species and species hybrids. By completing this proposal, I will connect variation in the gene regulatory network to molecular mechanisms that dictate cell size and shape, which will develop the posterior lobe as a premier system for systems-level studies of morphological development and evolution.