RUI: Collaborative Research: Adaptive Significance of Genomic Variation in a Montane Insect

随着地球气候变暖和多变，帮助生物体应对压力的特征将变得越来越重要。这项研究调查了生活在加利福尼亚州内华达山脉高海拔地区的甲虫，以研究编码处理能量和应对压力的蛋白质的基因变异如何影响新陈代谢和表现。这项研究试图发现氧气和温度的不同作用，前者是处理新陈代谢燃料所必需的，后者改变了这些过程发生的速度。它是基因组学、生理学和动物自然史的综合体，将提供一个独特的机会来理解有机体如何应对环境变化。它将为热塑性的进化提供洞察力，并可能揭示与温度和氧气压力相关的新基因。这项工作将增加两所以本科为主的大学的学生的研究机会。学生将接受实验设计和分析、提案和论文写作以及在科学会议上发表研究报告的培训，这将为他们提供现代工作环境下的市场技能。将为K12教育开发教育材料，说明如何使用基因组工具回答科学问题。该项目将在旧金山湾区和加利福尼亚州因约县附近的公共活动中进行推广活动。该项目将开发和使用基因组和转录工具，以获得对塞拉利昂柳叶甲适应温度和大气氧气的遗传基础的机械性见解。这项研究将检验这一假设，即线粒体和核基因组之间的相互作用对基因表达具有普遍影响，这种影响扩大到代谢能力和表现的差异。为了验证这一假设，我们将研究线粒体和核遗传背景不同的种群的幼虫的应激反应，这些种群发生在纬度和陡峭的海拔温度梯度上。幼虫将在一个高海拔的实验室中饲养，实验室中的氧气水平和温度将受到控制。在生长期末，将在这些不同条件下饲养的幼虫暴露在不同的温度和运行速度测量。通过对跑速分布上尾部和下部尾部的个体进行全基因组测序，将识别出与应激后跑步速度快和慢相关的遗传变异。对于这些相同的个体，将使用RNA测序来评估感兴趣基因的表达差异。随着自然温度或海拔梯度的变化，具有非同义单核苷酸多态(SNP)的感兴趣基因(例如，应激、缺氧、中枢代谢)将在全基因组测序实验中被识别，并用于构建可筛选更多个体的SNP小组。然后，上述饲养条件将被用来产生幼虫，并在其中检查代谢生理学。将测量代谢酶活性、线粒体呼吸、氧化损伤和耐热性，记录SNP变异，并使用定量聚合酶链式反应量化感兴趣基因的差异表达。综上所述，这项研究将揭示线粒体和核基因组如何在不断变化的环境中相互作用来应对压力。该项目将与来自瑞典斯德哥尔摩大学的世界领先的昆虫基因组学专家建立新的国际合作。国际合作者将帮助圣克拉拉大学和索诺马州立大学的本科生和硕士学生组织和领导为期两周的基因组学和生物信息学实践研讨会。此外，索诺马国立大学的硕士研究生将前往斯德哥尔摩大学接受基因组学和生物信息学方面的培训。国际活动和旅行将由NSF国际和综合活动办公室国际科学和工程部提供资金支持。

As the Earth's climate becomes warmer and more variable, characteristics that help organisms cope with stress will become increasingly important. This research investigates beetles living at high elevation in the Sierra Nevada mountains of California to study how variation in genes coding for proteins that process energy and respond to stress affect metabolism and performance. The research seeks to discover the contrasting roles of oxygen, which is necessary to process metabolic fuel, and temperature, which changes the rate at which these processes occur. It is a synthesis of genomics, physiology, and animal natural history that will provide a unique opportunity to understand how organisms cope with environmental change. It will provide insight into the evolution of thermal plasticity and may unveil novel genes associated with temperature and oxygen stress. The work will increase research opportunities for students at two primarily undergraduate universities. Students will be trained in experimental design and analysis, proposal and paper writing, and in the presentation of research at scientific meetings, which will provide them with marketable skills for the modern work environment. Educational materials will be developed for K12 education, which illustrate the use of genomic tools for answering scientific questions. Outreach events detailing the results of this project will occur at public events in the San Francisco Bay Area and near field sites in Inyo County, California.This project will develop and use genomic and transcriptomic tools to gain mechanistic insights into the genetic basis of adaptation to temperature and atmospheric oxygen in the Sierra willow leaf beetle Chrysomela aeneicollis. This research will test the hypothesis that interactions between mitochondrial and nuclear genomes have pervasive effects on gene expression that scale up to differences in metabolic capacity and performance. To test this hypothesis, the stress response will be investigated in larvae from populations that differ with respect to mitochondrial and nuclear genetic background and that occur along a latitudinal and steep altitudinal temperature gradients. Larvae will be reared at a high elevation laboratory in chambers where oxygen level and temperature will be manipulated. At the end of the growth period, larvae reared under these different conditions will be exposed to different temperatures and running speed measured. Genetic variation associated with fast and slow running speed after stress will be identified using whole genome sequencing of individuals in the upper and lower tails of the running speed distribution. Differences in expression of genes of interest will be assessed using RNA sequencing for these same individuals. Genes of interest (e.g. stress, hypoxia, central metabolism) that have non-synonymous single nucleotide polymorphism (SNP) variation along natural temperature or elevation gradients will be identified in whole genome sequencing experiments and used to build 'SNP panels' in which larger numbers of individuals can be screened. Rearing conditions described above will then be used to generate larvae in which metabolic physiology will be examined. Metabolic enzyme activity, mitochondrial respiration, oxidative damage and thermal tolerance will be measured, SNP variation recorded, and differential expression of genes of interest quantified using quantitative PCR. Taken together, this research will reveal how mitochondrial and nuclear genomes interact to cope with stress in a changing environment. The project will establish a new international collaboration with world leading experts in insect genomics from Stockholm University (Sweden). International collaborators will help organize and lead a two-week hands-on workshop on genomics and bioinformatics for undergraduate and Masters students from Santa Clara University and Sonoma State University. Additionally, Masters students from Sonoma State University will travel to Stockholm University to receive training in genomics and bioinformatics. International activities and travel will be supported by funds from the International Science and Engineering section of NSF's Office of International and Integrative Activities.