Investigating the processes of life in the cold: high resolution imaging of cellular proteostasis in Antarctic fish species

南极生态系统在一个孤立的环境中进化了数百万年。由于地质和气候因素的综合作用，南极是地球上最孤立的生态系统。这一受保护的环境已经稳定了1000多万年，在适应这一大陆方面产生了显著的多样性。在所有生态系统中，南极洲特有物种的比例最高，大约有17,000种海洋无脊椎动物。然而，南极气候现在正面临着前所未有的威胁。政府间气候变化专门委员会的《关注的五个理由》报告称，两极是气候变暖最快的地区。数百万年来一直保持稳定的生态系统及其预计的快速变化的气候的综合影响使南极适应寒冷的物种特别容易受到海洋气候变化的影响。这些物种的脆弱性在于其细胞机制的稳定性，特别是蛋白质的制造、折叠和维持。目前，人们对蛋白质在0摄氏度左右的稳定性机制知之甚少，而对南极鱼类等复杂生物的这种稳定性则知之甚少。为了了解寒冷中生命的基本机制，并将这种理解应用于南极生态系统即将发生的变化，英国南极调查局与剑桥大学的几个小组和英国的其他小组合作，开展了一项雄心勃勃、高度跨学科的项目。作为合作的一部分，该博士项目专门针对显微镜和荧光工具的开发，以研究南极物种的蛋白质生命周期。这个博士项目将专注于显微镜平台的开发（第一年）以及荧光报告方法（第二年）。的确，在低温下进行显微镜观察存在技术上的挑战。这包括适当的激光剂量以维持细胞中的生理条件，以及严格控制显微镜台的温度，以便在一系列条件下观察样品。我们也期待着与显微镜材料本身的物理特性相关的挑战（不同部分的收缩，折射率的变化……）。一旦显微镜本身适应极性条件，我们还必须开发在0℃左右操作的荧光工具。这些工具将用于研究影响蛋白质折叠和稳定性的因素，如粘度和细胞化学。将尝试两种测量细胞内粘度的方法，即单粒子跟踪（观察一个元素随时间的运动）和分子转子（根据细胞条件或快或慢地旋转）。我们将在寒冷中结合显微镜和荧光技术，将细胞表型与细胞（第3年）所经历的温度变化联系起来。这个项目是高度跨学科的，存在于生物学、物理学和工程学的交叉点，汇集了各种各样的科学技术。就EPSRC的战略和研究领域而言，该项目涉及以下领域：通过开发新的独特显微镜平台来分析科学，通过研究基本蛋白质折叠动力学来研究生物物理学和软物质物理学，或者通过开发蛋白质跟踪和温度和粘度测量来开发传感器和仪器

Antarctic ecosystems have evolved in a singularly isolated environment for millions of years. Due to a combination of geological and climatic factors, the Antarctic is the most isolated ecosystem on earth. This shielded environment has been stable for over 10 million years, giving rise to a remarkable diversity in adaptation to this continent. Antarctica has the highest proportion of endemic species of all ecosystems with around 17,000 marine invertebrate species. [1] Antarctic climate is however now facing an unprecedented threat. The Intergovernmental Panel on Climate Change's Five reasons for Concern [2] report the poles as being the fastest warming climates. The combined effects of an ecosystem that has been stable for millions of years and its projected rapidly changing climate make Antarctic cold-adapted species especially vulnerable to ocean climate change. The vulnerability of these species lies in the stability of their cellular machinery, and in particular the making, folding and maintaining of proteins. Very little is currently known about the mechanisms of protein stability around 0 C, and fewer still about this stability in complex organisms such as Antarctic fish. In order to understand the fundamental mechanisms of life in the cold, and to apply this understanding to incoming changes in Antarctic ecosystems, the British Antarctic Survey has partnered with several groups of the University of Cambridge and others in the UK in an ambitious and highly interdisciplinary project. As part of this collaboration, this PhD project is specifically aiming at the development of microscopy and fluorescence tools to study the protein lifecycle of Antarctic species. This PhD project will be focused on the development of a microscopy platform (year 1) along with fluorescence reporter methods (year 2). Indeed, microscopy at cold temperatures comes with technical challenges. These include the appropriate dosage of laser light to maintain physiological conditions in the cells, and a tight control of the microscope stage temperature so as to observe the samples across a range of conditions. We also expect challenges linked with the physics of the microscopy materials themselves (contraction of different parts, change in refractive indices... ). Once the microscope itself is adapted to polar conditions, we will also have to develop fluorescence tools operational around 0 C. These tools will be used to study factors affecting protein folding and stability, such as viscosity and cell chemistry. Two methods to measure intracellular viscosities will be tried, namely Single Particle Tracking (looking at the movement of one element over time) and Molecular Rotors (which rotate more or less quickly depending on cell conditions). We will use the combination of the microscopy and fluorescence techniques in the cold to link cellular phenotypes with the change of temperature experienced by the cell (year 3). This project is highly interdisciplinary and exists at the intersection of Biology, Physics and Engineering, bringing together a wide variety of scientific techniques. In terms of the EPSRC's strategies and research areas, this project touches on the following: Analytical sciences through the development of a new and unique microscopy platform, Biophysics and soft matter physics through the study of fundamental protein folding dynamics, or Sensors and Instrumentation through the development of protein tracking and temperature and viscosity measurements in