Collaborative Research: Characterizing the Constraints on Virus Infection of Cyanobacteria

“该奖项是根据2009年美国复苏和再投资法案（公法111-5）资助的。“众所周知，病毒在调节海洋生态系统的结构和功能方面发挥着重要作用。然而，我们缺乏细菌细胞受体如何抑制病毒感染的机制理解，这对海洋微生物食物网的生态学和进化动力学有影响。该项目将确定海洋蓝细菌Synechococcus sp. WH 7803外膜上三种已知噬藻体（podo-，myo-和siphoviridae）的受体。 研究人员将采用一系列经过验证的方法，使用恒化器生成病毒抗性聚球藻种群，使用监测表面附着的荧光病毒标签缩小突变体，并使用LC-MS/MS蛋白质组学分析表征抗性种群的外膜蛋白质互补。一旦受体被确定，现有数据库的生物信息学分析将与样本档案的分子生物学查询相结合，以记录海洋蓝藻基因组中不同受体的多样性和普遍性，以及自然种群表达的受体。恒化器中的实验室研究将检查受体表达的调节（使用荧光病毒和qRT-PCR进行基因表达），以便开始梳理蓝藻培养群体中病毒受体表达的环境控制。所有信息的收集方式将使其适合于生态竞争模型的分析，从而提供关于蓝细菌面临的抗感染成本的新见解，以及关于病毒感染细胞如何受到环境条件限制的新信息。该项目的更广泛影响包括在微生物学、分子生物学和蛋白质组学等跨学科领域培训新一代海洋科学家，在EPSCoR研究所（田纳西大学）和研究站（凯洛格生物站）的学生中，海洋地球化学和经典生态学。这项研究将为学生提供机会，与来自一个主要的联邦研究机构（橡树岭国家实验室）的研究人员一起工作，在那里可以获得大多数学术机构所没有的设备。这项研究的信息将直接纳入课程工作，教育和公共宣传，并通过与田纳西大学科学新闻计划的新伙伴关系传播。将招募一名或多名对科学新闻感兴趣的本科生，以制作适合K-12学生和公众的海洋微生物学和生态学纸质和电子传播材料。项目研究员将继续其招募代表性不足的群体和参加以方法为导向的课程的良好历史。这项研究产生的信息将是全新的，扩展和改变海洋微生物学和病毒生态学的基础，并深入了解全球气候和海洋生物化学的变化如何影响微生物的增殖和海洋病毒的控制。

"This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5)." It is well established that viruses play an important role in regulating the structure and function of marine ecosystems. However, we lack a mechanistic understanding of how bacteria cell receptors constrain virus infection, which has implications for the ecology and evolutionary dynamics of marine microbial food webs. This project will identify receptors for three known groups of cyanophage (podo-, myo- and siphoviridae) on the on the outer membrane of the marine cyanobacterium Synechococcus sp. WH7803. The researchers will employ a series of proven methods to generate virus-resistant Synechococcus populations using chemostats, to narrow the mutants using fluorescent virus tags that monitor surface attachment, and to characterize the outer membrane protein compliment of the resistant populations using LC-MS/MS proteomic analyses. Once the receptors are identified, bioinformatic analyses of existing databases will be coupled with molecular biological queries of sample archives to document the diversity and prevalence of the different receptors in marine cyanobacterial genomes as well as of those expressed by natural populations. Lab studies in chemostats swill examine the regulation of receptor expression (using fluorescence viruses and qRT-PCR for gene expression) so as to begin to tease apart the environmental controls on virus receptor expression in culture populations of cyanobacteria. All information will be collected in a manner that will make it suitable for analyses by ecological competition models, thereby providing new insights on the cost-of-resistance to infection that cyanobacteria face as well as new information on how virus infection of cells may be constrained by environmental conditions.Broader Impacts: The broader impacts of this project include the training of a new generation of marine scientists in interdisciplinary sciences of microbiology, molecular biology and proteomics, marine geochemistry and classical ecology of students at an EPSCoR Institution (The University of Tennessee) and a research station (Kellogg Biological Station). The study will provide opportunities for students to work with researchers from a major federal research facility (Oak Ridge National Laboratory) where access to equipment exists that is not available at most academic institutions. Information from this study will be directly incorporated into course work, educational and public outreach, and disseminated through a novel partnership with the Science Journalism program at the University of Tennessee. One or more undergraduates interested in science journalism will be recruited in order to generate materials for paper and electronic dissemination on marine microbiology and ecology that will be appropriate for K-12 students as well as the general public. The PIs will continue their well-established history of recruitment of underrepresented groups and participation in methods-oriented courses. Scientifically the information generated by this study will be completely novel, extending and transforming the foundations of marine microbiology and virus ecology as well as providing insight into how shifts in the global climate and ocean biogeochemistry might influence microbial proliferation and control by marine viruses.