EAGER: Investigate a novel way of light harvesting by marine phytoplankton: proton-pump rhodopsin in dinoflagellates

光合作用是海洋生态系统中唯一已知的光营养途径，直到21世纪初，在生活在阳光照耀的海洋表面的海洋细菌中发现了一种质子泵类型的视紫红质(蛋白紫质，或PR)，并证明了它在促进跨膜质子运输导致ATP产生方面的作用。自那以后，人们发现PR基因存在于海洋表层水域中13%至80%的海洋细菌和古生菌中。PIS最近在海洋甲藻中检测到一种与质子泵类型密切相关的视紫红质基因，但缺乏在这类生物中发挥质子泵作用的直接证据。鉴于甲藻作为海洋初级生产者和微型食草动物的重要性，应确定视紫红质光营养在维持甲藻生长方面是否发挥重要作用，使甲藻能够在营养有限的环境中胜过其他浮游植物。智力价值：这项研究将解决两个核心问题：1)在甲藻中发现的基因是否真的编码了一种吸收光线并导致产生ATP的视紫红质？2)这种基因在甲藻群落中有多常见？在讨论甲藻视紫红质对海洋生态系统的总能量获取有多大贡献之前，需要解决这些问题。本研究将：(1)将甲藻视紫红质基因转化到大肠杆菌细胞中，以在受控的实验室条件下测试光诱导的ATP生成；(2)分析长岛海湾有代表性的甲藻(养殖和野生)是否存在该基因；以及(3)检测在长岛海湾不同的温度(季节)和营养(空间)条件下，甲藻视紫红质序列的多样性。由于(没有质子泵功能)的风险和有限的背景工作，如果作为一个成熟的项目提交，这项提案不太可能成功。EIGHER计划提供了一种机制来探索这些有风险但潜在重要的问题。更广泛的影响：这项研究将为了解真核生物视紫红质可能对海洋光养有多大贡献提供必要的信息。这些结果将提供一个基础，以确定未来是否应该投入重大研究努力，以系统地调查该基因在海洋生态系统中的生态意义。该项目还将对教育和培训作出贡献。首先，研究生班(生物海洋学的分子方法)将参与该项目的一部分，作为课程的必修研究部分。第二，该项目将为一到两名本科生提供一般海洋科学研究，特别是浮游生物分子生态学的培训机会。

Photosynthesis was the only known pathway of phototrophy in marine ecosystems until the early 2000s, when a proton-pump type of rhodopsin (proteorhodopsin, or PR) was found to occur in marine bacteria living in the sunlit ocean surface, and its role in facilitating trans-membrane proton transport that leads to ATP production was demonstrated. It has since been found that the PR gene occurs in 13% to 80% of marine bacteria and archaea in the ocean's surface waters. The PIs recently detected a rhodopsin gene in marine dinoflagellates that is closely related to the proton-pump type, but direct evidence of its proton-pump action in this group of organisms is lacking. Given the importance of dinoflagellates as oceanic primary producers and micrograzers, it should be determined if rhodopsin-based phototrophy plays an important role in sustaining dinoflagellate growth, enabling dinoflagellates to outcompete other groups of phytoplankton in nutrient-limited environments. Intellectual Merit: This study will address two central questions: 1) Does the gene discovered in dinoflagellates indeed code for a rhodopsin that absorbs light and leads to the production of ATP? and 2) How common is this gene in dinoflagellate communities? These questions need to be addressed before moving on to the question of how much dinoflagellate rhodopsin contributes to overall energy acquisition in the marine ecosystem. The current study will: (1) transform the dinoflagellate rhodopsin gene into E. coli cells to test for light-induced ATP-generation under controlled laboratory conditions; (2) analyze representative dinoflagellates (cultured and wild) from Long Island Sound for presence of the gene; and (3) examine the diversity of dinoflagellate rhodopsin sequences under contrasting temperature (seasons) and nutrient (spatial) conditions in Long Island Sound. Due to the risk (of no proton pumping function) and limited background work, this proposal would not likely succeed if submitted as a full-fledged project. The EAGER program provides a mechanism for exploring these risky but potentially important questions. Broader Impacts: This study will provide essential information for understanding how much eukaryotic rhodopsin may contribute to marine phototrophy. The results will provide a basis on which to determine whether major research efforts should be invested in the future to systematically investigate the ecological significance of this gene in marine ecosystems. This project will also contribute to education and training. First, a graduate class (Molecular Approaches to Biological Oceanography) will be involved in part of the project as a required research component of the course. Second, the project will provide training opportunities to one or two undergraduate students in marine science research in general and molecular ecology of plankton in particular.