RoL:NSF-BSF: IMAGiNE: Temperature acclimation through RNA editing

创造有机体和维持其生命所需的所有信息都编码在它的DNA中。在任何给定时刻所需的信息部分在被解码为执行所需功能的蛋白质之前被临时复制到RNA中。操纵这种信息流的分子系统已经导致了现代生物学的一些最重要的工具，从基因克隆技术到CRISPR基因组工程。所有多细胞动物都可以在遗传信息通过RNA时对其进行编辑，但它们这样做的程度差别很大。鞘状头足类(鱿鱼、章鱼和墨鱼)编辑RNA的水平非常高，因此为研究这一过程提供了理想的模型。因为RNA是一种暂时的分子，所以RNA编辑是一个很好的工具，可以创造暂时的变化，帮助适应环境。这项拨款的工作旨在更好地了解头足类RNA编辑是如何用于温度适应的。利用最近开发的头足类基因敲除技术，产生高水平RNA编辑的酶(S)将被发现。此外，还将研究单个RNA编辑如何影响蛋白质。最后，将对生活在不同热环境中的相似鱿鱼物种的RNA编辑的进化基础进行比较。该项目将通过为波多黎各大学的学生举办关于基因组编辑的实践讲习班，加强对代表不足的少数群体的STEM教育。有关头足类生物学和RNA编辑的宣传将扩展到科学记者和犹太社区的犹太教士领袖。大多数生物体面临着多变的热环境，由于水的高导热性，水生外温体的温度波动尤其剧烈。所有的酶反应都受温度控制，但敏感度各不相同。协调不同温度下的多种酶反应是一项基本的生理挑战。该提案重点介绍了通过腺苷脱胺进行的RNA编辑以及如何将其用于温度驯化。所有多细胞后生动物都使用这一过程，但鞘类头足类动物使用最广泛。转录组范围内的屏幕已经发现了数万个RNA编辑站点，这些大型数据集提供了一个独特的机会，可以研究编辑是如何用于微调生理的，以及它是如何进化的。利用最近开发的头足类基因敲除方法，将探索高水平重新编码的酶基础，以及其温度敏感性的分子基础。此外，这项工作将研究温度敏感的编辑位点如何影响神经生理学中关键蛋白质的功能，并研究这些位点是如何在来自不同热环境的密切相关物种中选择的。该项目将加强对代表不足的少数群体的STEM教育，为头足类生物学创造资源，并提供多样化的社区外联。将为波多黎各大学的学生举办关于基因组编辑的讲习班。PIS将为头足类动物在研究中的人道和道德使用制定指导方针。在头足类生物和RNA编辑方面的推广将扩展到科学记者和犹太社区的犹太教士领袖。这项提议得到了综合组织系统部门的综合生态生理学项目，分子和细胞生物科学部门的遗传机制集群，以及生物科学局的生命规则项目的支持。该奖项反映了NSF的法定使命，并通过使用基金会的智力价值和更广泛的影响审查标准进行评估，被认为值得支持。

All the information required to create an organism, and for the organism to sustain its life, is encoded in its DNA. The portion of the information required at any given moment is temporarily copied into RNA before being decoded into the proteins that execute required functions. Molecular systems that manipulate this flow of information have led to some of the most consequential tools for modern biology, from gene cloning technologies to CRISPR genome engineering. All multicellular animals can edit genetic information as it passes through RNA, however the extent to which they do so varies widely. The coleoid cephalopods (squid, octopus, and cuttlefish) edit RNA at exceptionally high levels, and thus provide ideal models to study the process. Because RNA is a transient molecule, RNA editing is a good tool to create temporary changes that help with environmental acclimation. The work in this grant seeks to better understand how cephalopod RNA editing is used for temperature acclimation. Using recently developed cephalopod gene knockout technologies, the enzyme(s) that generate high-level RNA editing will be uncovered. In addition, how individual RNA edits affect proteins will be studied. Finally, the evolutionary underpinnings of RNA editing will be compared between similar squid species that inhabit different thermal environments. This project will enhance STEM education for underrepresented minorities through hands-on workshops on genome editing given to students from the University of Puerto Rico. Outreach on cephalopod biology and RNA editing will be extended to science journalists, and rabbinic leaders from Jewish community.Most organisms are confronted by a variable thermal environment and temperatures fluctuations are particularly acute for aquatic ectotherms due to the high thermal conductivity of water. All enzymatic reactions are governed by temperature, but the degree of sensitivity varies. Coordinating multiple enzymatic reactions across temperatures is a fundamental physiological challenge. This proposal focuses on RNA editing through adenosine deamination and how it is used for temperature acclimation. All multicellular metazoans use this process, however the coleoid cephalopods use it most extensively. Transcriptome-wide screens have uncovered tens of thousands of RNA editing sites, and these large data sets provide a unique opportunity to examine how editing is used to fine-tune physiology, and how it evolves. Taking advantage of recently developed gene knockout methods for cephalopods, the enzymatic underpinnings of high-level recoding will be explored, as well as the molecular basis of its temperature sensitivity. In addition, the work will investigate how temperature sensitive editing sites affect the function of key proteins in neurophysiology and examine how these sites are selected in closely related species from different thermal environments. This project will enhance STEM education for underrepresented minorities, generate resources for cephalopod biology, and provide diverse community outreach. Workshops on genome editing will be given to students from the University of Puerto Rico. The PIs will create guidelines for the humane and ethical use of cephalopods in research. Outreach on cephalopod biology and RNA editing will be extended to science journalists, and rabbinic leaders from Jewish community.This proposal was supported by the Integrative Ecological Physiology Program within the Division of Integrative Organismal Systems, the Genetic Mechanisms Cluster within the Division of Molecular and Cellular Biosciences, and Rules of Life Program within the Directorate for Biological Sciences.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.