CAREER: The Molecular Mechanisms of Cold-Induced Mitochondrial Biogenesis

温度对动物的生理有深远的影响。温度降低时发生的一种变化是某些组织中线粒体密度的增加。这个过程对于维持动物的有氧代谢能力至关重要，但关于它是如何被调节的，以及为什么它发生在一些组织中，而不是其他组织中，问题仍然存在。本项目将以三刺棘鱼（Gasterosteus aculetaus）为模式生物来解决这些问题。这项研究将验证活性氧（ROS）是由线粒体在低温下产生的，并且是诱导控制线粒体生物发生的基因表达所必需的假设。膜流动性随着温度下降而降低，并可能破坏线粒体电子传递链蛋白的活性，导致ROS的形成。这些ROS可能直接调节线粒体生物发生基因的表达，或者，高水平的ROS可能刺激一氧化氮合酶的活性，一氧化氮合酶产生一氧化氮并触发线粒体生物发生。这两种可能性都将受到考验。刺鱼将在实验室中保持在20摄氏度，然后转移到5摄氏度，并保持6周。在整个驯化期间，将采集动物，测量ROS水平、线粒体蛋白、基因表达和一氧化氮合酶活性。这些参数将在几种组织类型中进行测量，包括那些经历线粒体生物发生的组织（氧化骨骼肌）和那些可能没有的组织（肝脏）。不同组织类型对低温反应的差异可能阐明线粒体生物发生的组织特异性机制。如果ROS和/或NO水平在冷驯化的早期阶段增加，那么动物将使用ROS和/或NO形成抑制剂进行处理。如果需要这些自由基中的任何一种来诱导线粒体生物发生，那么抑制它们的产生应该可以防止线粒体密度在低温下的增加。这项工作的结果将导致对线粒体生物发生的分子机制有更深入的了解。这是一个至关重要的过程，对所有生物体的健康至关重要。这项研究将通过与阿拉斯加大学阿拉斯加土著科学与工程项目的合作，涉及阿拉斯加土著学生的本科生和研究生。这些学生还将帮助在当地一所农村学校开展一项推广计划，让学生学习动物在低温下生存的各种策略，重点是生理、细胞和分子适应。学生将学习动物的重要作用，甚至是那些在当地环境中发现的动物，以解决生物学的基本问题。

Temperature has a profound effect on the physiology of animals. One modification that occurs in response to a decrease in temperature is an increase in mitochondrial density in some tissues. This process is critical for maintaining aerobic metabolic capacity in animals, yet questions remain about how it is regulated, and why it occurs in some tissues, yet not others. This project will address these questions, using the threespine stickleback Gasterosteus aculetaus as a model organism. This investigation will test the hypothesis that reactive oxygen species (ROS) are produced by mitochondria in response to cold temperature and are required to induce the expression of genes controlling mitochondrial biogenesis. Membrane fluidity decreases as temperature declines and may disrupt the activity of the electron transport chain proteins of the mitochondrion, leading to the formation of ROS. These ROS may directly regulate mitochondrial-biogenic gene expression, or alternatively, high levels of ROS may stimulate the activity of nitric oxide synthase which produces nitric oxide and triggers mitochondrial biogenesis. Both of these possibilities will be tested. Sticklebacks will be maintained at 20 degrees C in the laboratory and then shifted to 5 degrees C and held over a period of six weeks. Throughout the acclimation period, animals will be harvested for measuring levels of ROS, mitochondrial proteins, gene expression, and nitric oxide synthase activity. These parameters will be measured in several tissue types, including those that undergo mitochondrial biogenesis (oxidative skeletal muscle), and those that may not (liver). The differences in response to cold temperature among different tissue types may elucidate the mechanisms underlying the tissue specificity of mitochondrial biogenesis. If ROS and/or NO levels increase during the early stages of cold acclimation, then animals will be treated with inhibitors of ROS and/or NO formation. If either of these free radical species are required to induce mitochondrial biogenesis, then inhibiting their production should prevent increases in mitochondrial density in response to cold temperature. The results from this work will lead to a greater understanding of the molecular mechanisms governing mitochondrial biogenesis. This is a vital process, essential to the health of all organisms. This research will involve undergraduate and graduate Alaska Native students through a collaboration with the Alaska Native Science and Engineering Program at the University of Alaska. These students will also help to develop an outreach program at a local rural school which engages students in learning about the variety of strategies that animals use to survive at cold temperature, with an emphasis on physiological, cellular and molecular adaptations. Students will learn the important role of animals, even those found in local environments, for addressing fundamental questions in biology.