Feeding responses of marine phagotrophic protists: cell biological mechanisms

摄食原生生物作为细菌和浮游植物生物量的主要消费者和再矿化者，在海洋中上层食物网中发挥着核心作用。原生动物食草动物可以根据猎物细胞的特征表现出复杂的摄食反应，包括选择性摄食或回避。这种选择性有可能影响海洋中的碳循环。阐明海洋捕食性原生动物选择性地以不同类型的猎物为目标的生化机制对于了解海洋食物网的结构和功能至关重要。研究人员的初步结果表明，原生动物的摄食行为，包括化学感觉反应和猎物摄取，实际上涉及基本的细胞生物学机制：即配体化合物与与细胞膜相关的受体分子结合，激活细胞内信号转导途径，导致细胞生化和行为的改变。他们将研究信号化合物与原生生物细胞膜上的受体分子结合的作用，以及随后的信号转导级联反应在控制原生生物对猎物的化学感觉反应和猎物摄取方面的作用。这个项目将包括1)用已知的和最近分离的海洋原生生物物种进行实验，然后2)用从俄勒冈州沿海海水中生长的原生生物组合进行实验。这项研究将集中于确定配体结合和信号转导过程在多大程度上控制原生动物的取食行为，以及细胞-表面配体-受体结合的差异在多大程度上可以解释不同类型猎物之间的选择性。该方法将包括：1)药理学方法：由糖和凝集素组成的糖链微阵列，以测试原生生物和猎物细胞膜上是否存在特定的糖-凝集素结合部位，并使用特定蛋白激酶、G蛋白和G蛋白连接受体的选择性抑制剂来测试信号转导通路在海洋原生动物取食行为中的作用，以及2)旨在表明配体-凝集素结合的特异性，以及细胞内信号转导机制，在原生生物分离物和异养鞭毛的原位组合中，对猎物的选择性/切换和捕食率具有重要作用。智力优势：原生生物摄食反应的生化机制可能是理解海洋系统中微生物种群的这种自上而下的主要控制的基础。这项研究调整了药理学/细胞生物学中使用的方法，以一种可能导致与海洋微生物生态相关的新发现的方式，以及在评估原生动物食草动物就地影响的新方法中。更广泛的影响：该项目将支持一名研究生的博士论文研究，并将包括一名本科生参与实验。该项目的成果将通过演讲、出版物、讲座和调查人员网站进行传播。

Phagotrophic protists play a central role in marine pelagic food webs as major consumers and remineralizers of bacterial and phytoplankton biomass. Protistan grazers can show complex feeding responses, including selective ingestion or avoidance, based on characteristics of prey cells. Such selectivity has the potential to affect carbon cycling in the sea. Elucidating the biochemical mechanisms by which marine phagotrophic protists might selectively target distinct types of prey is vital to understanding the structure and function of marine food webs. The investigators have shown preliminary results which show that protistan feeding behavior, including chemosensory response and prey ingestion, does in fact involve basic cell biological mechanisms: i.e. binding of ligand compounds to receptor molecules associated with cell membranes, which activates intracellular signal transduction pathways leading to changes in cell biochemistry and behavior. They will conduct an investigation of the role of binding of signal compounds to receptor molecules on protistan cell membranes, and of the consequent signal transduction cascades in controlling protistan chemosensory response to prey, and prey ingestion. This project will involve 1) experiments with both known and recently isolated species of marine protists, followed by 2) experiments with protist assemblages grown up from Oregon coastal seawater. The research will focus on determination of the extent to which ligand binding and signal transduction processes control protist feeding behavior, and the extent to which differences in cell-surface ligand-receptor binding can explain selectivity between alternate types of prey. The approach will include: 1) pharmacological methods: glycomic microarrays composed of sugars and lectins to test for the presence of specific sugar-lectin binding sites on protist and prey cell membranes, and use of selective inhibitors of specific protein kinases, G proteins, and G protein-linked receptors to test for the role of signal transduction pathways in the feeding behavior of marine protists, and 2) experiments designed to show whether specificity of ligand-lectin binding, and whether intracellular signal transduction mechanisms, have a significant role in prey selectivity/switching and rates of prey ingestion by protist isolates and by in situ assemblages of heterotrophic flagellates.Intellectual merit: The biochemical mechanisms underlying protistan feeding response are likely to be fundamental to understanding this major top-down control of microbial populations in marine systems. This study adapts approaches used in pharmacology/cell biology in a way that could result in new discoveries relevant to marine microbial ecology, and in novel approaches to assessing the in situ impact of protist grazers. Broader impacts: This project will support the PhD thesis research of a graduate student and will also involve an undergraduate student in the experiments. Results of the project will be disseminated through presentations, publications, lectures, and the investigators' website.