Genetically engineered ants to label and study neurons involved in social behavior

摘要 这项提议的目标是为调节社会活动的神经元回路产生遗传报告细胞系。 行为Harpegnathos saltator蚂蚁，并利用这些线可视化脑重塑作为成年工人 重新编程让自己像蚁后一样行动 社会性昆虫是研究大脑功能和行为的表观遗传调控的新兴模式系统 因为相同的基因组在不同的种姓中指定了不同的行为状态。大脑是否以及如何 工蚁和蚁后的不同连接方式实现了不同的种姓特定行为集，这是未知的。 在Harpegnathos蚂蚁中，成年工人可以通过种姓转换成为女王， 生殖、生理和行为的表型变化。Harpegnathos蚂蚁的这一特征使得 基因工程等位基因的容易繁殖，因为任何个体都可以繁殖。 成年羽颚龙的工蜂成为蚁后的能力为机械化的 成人大脑可塑性的解剖单细胞RNA-seq分析揭示了细胞内的主要变化， 影响神经元和神经胶质的组成，表明大脑的结构重塑伴随着 种姓的转变。指导这种脑重塑的分子信号仍然未知。 我们之前已经证明，神经肽corazonin刺激狩猎，一个工人特有的行为， 随着工蚁成为蚁后，我们已经获得了新的初步证据， 人血管加压素的同系物以种姓特异性方式表达，并且可能也调节一个子集 的社会行为。我们假设这些神经肽的信号引导大脑 重塑事件，巩固社会行为的转换在种姓过渡。 在目标1中，我们将利用转基因方法来标记产生corazonin和加压素的神经元， 他们的投影与膜结合的绿色荧光蛋白，并观察其重塑过程中的自然种姓过渡。 在目标2中，我们将利用CRISPR/Cas9产生敲入驱动和报告细胞系来标记神经元， 反应corazonin或加压素和可视化他们的可塑性在细胞和突触水平的反应 神经肽的作用。 这些实验将揭示Harpegnathos蚂蚁成年大脑可塑性的关键信息，并提供 复杂的遗传工具，进一步解剖这些社会行为的表观遗传调控， 蚂蚁考虑到corazonin和加压素在哺乳动物中有对应物，我们的研究结果有望对哺乳动物产生影响。 对我们理解神经肽如何调节大脑可塑性和社会行为产生了广泛的影响。

ABSTRACT The goal of this proposal is to generate genetic reporter lines for neuronal circuits that regulate social behavior in Harpegnathos saltator ants and to utilize these lines to visualize brain remodeling as adult workers reprogram themselves to behave as ant queens. Social insects are an emerging model system to study the epigenetic regulation of brain function and behavior because the same genome specifies alternative behavioral states in different castes. Whether and how brains of workers and queens are differently wired to implement distinct sets of caste-specific behaviors is not known. In Harpegnathos ants, adult workers can become queens via a caste transition that involves dramatic phenotypic changes in reproduction, physiology, and behavior. This feature of Harpegnathos ants allows for easy propagation of engineered alleles, as any individual can become reproductive. The ability of adult Harpegnathos workers to become queens offers unique opportunities for the mechanistic dissection of adult brain plasticity. Single-cell RNA-seq analyses revealed major changes in cellular composition affecting both neurons and glia, indicating that a structural remodeling of the brain accompanies the caste transition. The molecular signals that direct this brain remodeling remain unknown. We previously showed that the neuropeptide corazonin stimulates hunting, a worker-specific behavior, and is downregulated as workers become queens. We have since obtained new preliminary evidence that the ant homolog of human vasopressin is expressed in a caste-specific manner and likely also regulates a subset of social behaviors in Harpegnathos ants. We hypothesize that signaling by these neuropeptides directs brain remodeling events that underpin the switch in social behaviors during the caste transitions. In Aim 1, we will utilize a transgenic approach to label corazonin- and vasopressin-producing neurons and their projection with membrane-bound GFP and to observe their remodeling during the natural caste transition. In Aim 2, we will utilize CRISPR/Cas9 to generate knock-in driver and reporter lines to label neurons that respond to corazonin or vasopressin and visualize their plasticity at the cellular and synaptic level in response to the neuropeptides. These experiments will reveal crucial information on adult brain plasticity in Harpegnathos ants and provide sophisticated genetic tools for the further dissection of the epigenetic regulation of social behavior in these ants. Given that corazonin and vasopressin have mammalian counterparts, our results are expected to have a broad impact on our understanding of how neuropeptides regulate brain plasticity and social behavior.