RUI: Characterization of the gene regulatory network promoting hormogonium development in the filamentous cyanobacterium Nostoc punctiforme

丝状蓝藻是一组光合微生物，大约在25亿年前首次进化，是地球上最早的多细胞生物之一。这些相对简单的多细胞生物为研究发育生物学的基本方面提供了一个极好的、具有成本效益的模型系统。此外，由于这些细菌相对容易进行基因改造，并且能够将阳光转化为能量并为某些植物提供氮基肥料的来源，因此它们对生物燃料生产和农业的发展具有相当大的兴趣。这个项目的重点是表征遗传机制控制的特殊细丝的发展，能够运动，使用模式菌株Nostoc点状。作为更广泛影响的一部分，该项目还将为本科生和研究生提供最先进的遗传和分子技术培训，并向当地学校提供外展计划，这些学校为不同的学生群体提供服务，其中许多学生经济上处于不利地位。该项目的一些方面将被纳入教学实验室，一些本科生和研究生将参与研究，其中包括从太平洋大学社区参与计划（c.i.p）招募的学生，该计划为来自种族代表性不足和经济条件不利背景的第一代大学生提供服务。从cip招募的学生将获得暑期研究助教的支持，并参加当地学校5 -8年级教室的外展计划，在那里他们将传播使用便携式荧光显微镜探索细胞生物学的学习模块。丝状蓝藻是发育最复杂的原核生物之一，对这些生物的研究为发育生物学提供了有价值的见解。许多物种发育出一系列特殊的细胞类型，包括被称为原丝的活动丝，以促进扩散。藻殖菌对于细胞上结构的形成和固氮植物和真菌-蓝藻共生的建立也是必不可少的，这对全球固氮有重要贡献。目前，潜在的分子相互作用，控制均衡发育和生态重要的均衡依赖过程在很大程度上是不明确的。基于最近的研究和初步数据，工作假设是同源性的发育是由一个保守的基因调控网络控制的，包括sigma因子、伴侣转换调控系统、hmp趋化系统和一些其他未表征的成分。本项目有两个具体目标：1)识别和表征促进同源性生殖发育的基因调控网络的组成部分；2)明确sigma因子和伴侣转换调控系统在同源性生殖发育中的作用。该项目采用正向和反向遗传方法，结合一系列突变菌株的表型分析，包括形态分析和赤藻特异性蛋白质和多糖表达的免疫学分析，以及转录组学研究，以确定促进模型丝状蓝藻中赤藻发育的基因调控网络。该项目的完成将为促进藻殖菌发育的基因调控网络模型奠定基础，该模型可用于合理设计丝状蓝藻的转基因菌株，用于生物燃料生产和新型植物-蓝藻共生等各种应用。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

The filamentous cyanobacteria are a group of photosynthetic microorganisms that first evolved about 2.5 billion years ago, making them one of the first multicellular organisms on earth. These relatively simple multicellular organisms provide an excellent, cost-effective model system for studying fundamental aspects of developmental biology. Additionally, because these bacteria are relatively easy to modify genetically, and are capable of both converting sunlight into energy and providing a source of nitrogen-based fertilizer to certain plants, they are of considerable interest for development in biofuel production and agriculture. This project is focused on characterizing the genetic mechanism controlling the development of specialized filaments that are capable of movement, using the model strain Nostoc punctiforme. As part of the broader impacts, this project will also provide training for both undergraduate and graduate students in state-of-the art genetic and molecular techniques, as well as an outreach program to local area schools which serve a diverse student body, many of whom are economically disadvantaged. Aspects of this project will be incorporated into teaching labs and several undergraduate and graduate students will participate in the research, including students recruited from the Community Involvement Program (C.I.P.) at University of the Pacific, which serves first generation college students from ethnically underrepresented and economically disadvantaged backgrounds. Students recruited from C.I.P. will be provided support for summer research assistantships and also participate in an outreach program for 5th-8th grade classrooms at local schools where they will disseminate a learning module using a portable fluorescence microscope to explore cell biology.Filamentous cyanobacteria are among the most developmentally complex prokaryotes and the study of these organisms has provided valuable insight into developmental biology. Many species develop an array of specialized cell types including motile filaments termed hormogonia that facilitate dispersal. Hormogonia are also essential for the formation of supracellular structures and the establishment of nitrogen-fixing plant- and fungal-cyanobacterial symbioses that contribute significantly to global nitrogen fixation. Currently, the underlying molecular interactions controlling hormogonium development and ecologically important hormogonium-dependent processes are largely undefined. Based on recent studies and preliminary data, the working hypothesis is that hormogonium development is governed by a conserved gene regulatory network that includes sigma factors, partner-switching regulatory systems, the hmp chemotaxis system, and several additional uncharacterized components. This project has two specific aims: 1) Identify and characterize components of the gene regulatory network promoting hormogonium development, and 2) Define the role of sigma factors and partner-switching regulatory systems in hormogonium development. The project uses forward and reverse genetic approaches combined with a range of phenotypic analyses of mutant strains including morphological analyses and immunological assays of hormogonium-specific protein and polysaccharide expression, as well as transcriptomic studies to define the gene regulatory network promoting hormogonium development in the model filamentous cyanobacterium Nostoc punctiforme. Completion of this project will lay the foundation for a model of the gene regulatory network promoting hormogonium development that can then be used to rationally design genetically modified strains of filamentous cyanobacteria for various applications including biofuel production and novel plant-cyanobacterial symbioses.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.