Studies of Antifreeze Proteins and Related Overwintering Adaptations in Arctic and Anarctic Insects

北极和南极昆虫的抗冻蛋白及相关越冬适应的研究

• 批准号: 0352919
• 负责人: Brian Barnes
• 金额: $ 11.5万
• 依托单位: University of Alaska Fairbanks Campus
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-06-01 至 2006-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0352919&HistoricalAwards=false
• 关键词: Studies; Antifreeze; Proteins; Related; Overwintering

This is a collaborative proposal by Principal Investigators at the University of Alaska-Fairbanks and Purdue University. Antifreeze proteins (AFPs) are found in a number of diverse organisms including fish, insects, spiders, mites, plant, and bacteria. While there is tremendous structural variation among these AFPs they all appear to lower the freezing point of water by a similar non-colligative mechanism. However, the insect AFPs are, by far, the most active, generally functioning to prevent freezing in freeze susceptible insects by inhibiting ice nucleating agents and innoculative freezing across the cuticle. However, a few AFP-producing insects are freeze tolerant and the function of AFPs in these is unknown. Two insect AFPs have been characterized. One type has been described from two species of beetles, but there is variation in sequence, etc., both within and between the species. Until recent research identified 18 species with AFPs, these proteins had not been reported in Alaskan or arctic insects. Certain of these have been chosen for further study, presenting the opportunity to extend understanding of insect AFPs to extremely cold tolerant species, and in the case of the beetle Cucujus clavipes ,to compare populations from arctic Alaska to those from Indiana. The mean winter supercooling point of Alaskan Cucujus larvae are -42oC, with some individuals supercooling to -57oC, while that of Indiana larvae is -24oC. AFP activity in Alaskan Cucujus is, by far, the highest ever described, even though the AFPs are similar to those of other beetles. Understanding the structure/function relationships of the Cucujus AFPs, and the enhancers (other proteins and polyols) which increase their activity, is of prime concern. In addition to AFPs, a suite of other overwintering adaptations (extreme dehydration to 1/3 normal body water, multimolar glycerol concentrations, diapause) contribute to the low supercooling points of Alaskan Cucujus. These are being studied, and using microarray technology and other means to monitor molecular markers, the Principal Investigators will attempt to understand the integration of these adaptations. Additional investigations involve the structure/function relationships of three other Alaskan beetle AFPs, and that of the insect type of AFP. Two of the beetle species are freeze tolerant and this is the first study of AFPs from freeze tolerant insects. The Principal Investigators will continue to screen additional species for the presence of AFPs (to identify interesting candidates for future study), and to monitor microhabitat temperatures and overwintering mortality of select species. The broader impact of this study concerns its effects on 1) biological education and 2) applications of AFPs in science and technology. Post-docs, PhD and undergraduate students, and high school teachers will work on this project. The Principal Investigators believe that the integrated training (in modern molecular techniques such as microarray, field biology, physiology of cold tolerance, protein biochemistry, etc.) at they will receive provides critical preparation for success in today's highlyintegrative and changing research environment. This will provide a direct benefit to teaching in biology. In addition to research papers, the Principal Investigators will incorporate these studies in review articles, symposium presentations. Also, as they are the most active AFPs known, insect AFPs (especially from Alaskan beetles) have tremendous potential for applied studies. For example, they have generated transgenic plants producing insect AFPs which lower the plant freezing temperature. Similar work using Alaskan beetle AFPs should be even more productive.

这是阿拉斯加大学费尔班克斯分校和普渡大学的首席研究员的合作提案。 抗冻蛋白 (AFP) 存在于许多不同的生物体中，包括鱼、昆虫、蜘蛛、螨虫、植物和细菌。虽然这些 AFP 之间存在巨大的结构差异，但它们似乎都通过类似的非依数机制来降低水的冰点。然而，迄今为止，昆虫 AFP 是最活跃的，通常通过抑制冰成核剂和穿过角质层的接种冷冻来防止易受冻害的昆虫受冻。 然而，一些产生 AFP 的昆虫具有耐冻性，并且 AFP 在这些昆虫中的功能尚不清楚。两种昆虫 AFP 已得到表征。已从两种甲虫中描述了一种类型，但在物种内部和物种之间存在序列等差异。 在最近的研究确定 18 个物种具有 AFP 之前，这些蛋白质在阿拉斯加或北极昆虫中尚未有报道。其中某些已被选择进行进一步研究，为将昆虫 AFP 的理解扩展到极冷物种提供了机会，并且以甲虫 Cucujus clavipes 为例，将北极阿拉斯加的种群与印第安纳州的种群进行比较。阿拉斯加杜鹃幼虫的平均冬季过冷点为-42℃，有些个体过冷至-57℃，而印第安纳幼虫的冬季过冷点为-24℃。尽管 AFP 与其他甲虫相似，但迄今为止，阿拉斯加 Cucujus 中的 AFP 活性是最高的。了解 Cucujus AFP 的结构/功能关系以及增强其活性的增强剂（其他蛋白质和多元醇）是首要关注的问题。除了 AFP 之外，一系列其他越冬适应（极度脱水至正常体内水分的 1/3、多摩尔甘油浓度、滞育）也导致了阿拉斯加黄瓜的过冷点较低。这些正在研究中，并使用微阵列技术和其他手段来监测分子标记，主要研究人员将尝试了解这些适应的整合。其他研究涉及其他三种阿拉斯加甲虫 AFP 以及昆虫类型 AFP 的结构/功能关系。其中两种甲虫具有耐冻性，这是首次对耐冻昆虫的 AFP 进行研究。 首席研究员将继续筛选其他物种是否存在 AFP（以确定未来研究中感兴趣的候选者），并监测选定物种的微生境温度和越冬死亡率。 这项研究更广泛的影响涉及其对 1) 生物教育和 2) AFP 在科学技术中的应用的影响。 博士后、博士生、本科生以及高中教师将参与该项目。 首席研究员相信，他们将接受的综合培训（现代分子技术，如微阵列、现场生物学、耐冷生理学、蛋白质生物化学等）为他们在当今高度综合和不断变化的研究环境中取得成功提供了关键的准备。这将为生物学教学带来直接的好处。除了研究论文外，主要研究人员还将这些研究纳入评论文章、研讨会报告中。 此外，由于昆虫 AFP（尤其是来自阿拉斯加甲虫）是已知最活跃的 AFP，因此具有巨大的应用研究潜力。例如，他们已经培育出产生昆虫 AFP 的转基因植物，从而降低植物的冷冻温度。使用阿拉斯加甲虫 AFP 进行的类似工作应该会更有成效。