Collaborative Research: From phylogeny to biomolecules: a cross-scale approach to understand the making of a unique carbon-concentrating mechanism in hornworts

合作研究：从系统发育到生物分子：一种跨尺度的方法来了解金鱼藻独特的碳浓缩机制的形成

• 批准号: 2213841
• 负责人: Fay-Wei Li
• 金额: $ 50.37万
• 依托单位: Boyce Thompson Institute Plant Research
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2213841&HistoricalAwards=false
• 关键词: Collaborative; Research; phylogeny; biomolecules; cross

Nature’s carbon fixing enzyme, Rubisco, is notoriously inefficient - often limiting the growth rate of photosynthetic organisms, including crop species. Some organisms have found a way around this problem by packaging Rubisco into compartments, called pyrenoids, which concentrate substrate CO2 around Rubisco active sites. Hornworts are the only land plants that have a pyrenoid. This project seeks to identify the hornwort pyrenoid components, characterize the hornwort carbon-fixing enzyme (i.e. Rubisco) housed within the pyrenoids, and determine how the various pyrenoid components interact with each other. The results of this study will provide evolutionary, biochemical, and mechanistic insights into the hornwort pyrenoid, and make a first step towards gathering the requisite understanding to transplant hornwort pyrenoids into crop plants to boost carbon fixation. This project will also train diverse undergraduates in a range of computational and experimental techniques, including plant tissue culture and transformation and bioinformatics. In addition, a three-part video series will be produced to provide a fun and informative introduction to hornwort diversity, evolution and pyrenoids, and distributed on YouTube and other online platforms. The importance of hornwort and Rubisco research will be further promoted through established podcast series.Photosynthesis is the source of energy for almost every life on earth, and is often limited by the carbon fixing enzyme, Rubisco. Some organisms have evolved ingenious ways to circumvent these limitations, such as pyrenoids, which are a kind of membrane-less organelle that liquid-liquid phase separates Rubisco from the stroma. Pyrenoids actively concentrate substrate CO2 around Rubisco active sites, thereby greatly enhancing the efficiency of carbon fixation. Hornworts are the only land plants with a pyrenoid, and these have translational potential for improving C3 crop efficiency. The fact that hornwort pyrenoids have been gained and lost over the course of evolution offers a unique phylogenetic replication for comparative studies. Putative pyrenoid components will be identified using comparative genomics, RNA-seq, and proteomics, while leveraging repeated pyrenoid-present/absent transitions. The subcellular localization of putative pyrenoid components and Rubisco subunits will be validated using in vivo localization approaches, and Rubisco populations structurally characterized. A constructionist biology approach will identify and characterize interactions between Rubisco and recombinantly-produced putative pyrenoid components in vitro, and will be used to assess the potential for Rubisco phase separation.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.

众所周知，自然界的碳固定酶Rubisco效率低下，常常限制光合生物的生长速度，包括农作物。一些生物已经找到了一种解决这个问题的方法，将Rubisco包装到称为蛋白核的隔间中，这些隔间将底物CO2集中在Rubisco活性位点周围。金鱼藻是唯一的陆地植物，有一个核。该项目旨在确定金鱼藻类蛋白核组分，表征蛋白核内的金鱼藻固碳酶（即Rubisco），并确定各种蛋白核组分如何相互作用。这项研究的结果将为金鱼藻蛋白核提供进化、生物化学和机制方面的见解，并为收集必要的理解将金鱼藻蛋白核移植到作物中以促进碳固定迈出第一步。该项目还将在一系列计算和实验技术方面培训不同的本科生，包括植物组织培养和转化以及生物信息学。此外，还将制作一个由三部分组成的视频系列，以提供关于金鱼藻多样性、进化和蛋白核的有趣和翔实的介绍，并在YouTube和其他在线平台上分发。通过已建立的播客系列，将进一步宣传金鱼藻和Rubisco研究的重要性。光合作用是地球上几乎所有生命的能量来源，但往往受到碳固定酶Rubisco的限制。一些生物已经进化出巧妙的方法来规避这些限制，例如蛋白核，这是一种无膜细胞器，将Rubisco与基质液-液相分离。类核蛋白主动地将底物CO2集中在Rubisco活性位点周围，从而大大提高了碳固定的效率。金鱼藻是唯一具有蛋白核的陆地植物，这些植物具有提高C3作物效率的转化潜力。事实上，金鱼藻蛋白核在进化过程中获得和失去了比较研究提供了一个独特的系统发育复制。假定的蛋白核组分将使用比较基因组学，RNA-seq和蛋白质组学来鉴定，同时利用重复的蛋白核存在/不存在的转换。将使用体内定位方法验证假定的蛋白核成分和Rubisco亚基的亚细胞定位，并对Rubisco群体进行结构表征。一个建构主义的生物学方法将确定和表征Rubisco和重组产生的假定蛋白核成分之间的相互作用在体外，并将被用来评估Rubisco相分离的潜力。这个奖项反映了NSF的法定使命，并已被认为是值得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的支持。