Genes and their products for low temperature stress resistance

在一些越冬生物，如某些鱼、植物和昆虫中，低温下的生存通过一些基因的表达而得到提高，其中包括那些独特的抗冻蛋白(AFP)。AFPS与形成的冰晶结合在一起，这种斑驳的覆盖物阻止了冰的生长。在易受冰冻影响的物种中，当温度进一步下降时，AFP不再“容纳”冰晶，随后出现爆炸性增长，导致冰冻，然后死亡。相比之下，耐冻物种在较高的零下温度下冻结，AFP使单个冰晶保持较小，并提供细胞保护。具有讽刺意味的是，考虑到关于AFP的丰富的生化数据，这些蛋白质对冷冻生存的实际贡献尚未确定。我们现在是第一个做到这一点的人，我们生产出了带有AFP的转基因雀麦草。我们必须优化这个系统，以便更充分地探索这些蛋白质的重要性。我们的研究表明，来自耐冻草的AFP(f-tAFP)有助于形成一个由低温、病原体和免疫相互作用组成的复杂网络。因此，我们推测f-tAFP具有多种作用：(I)通过组织冰来保护细胞的完整性，(Ii)‘破坏’细菌对冰的成核，以及(Iii)对耐低温细菌的更广泛的免疫反应。创新地，我们预计耐冻昆虫和微生物群落将表明，其他f-tAFP也是作为免疫反应的一部分而进化的。然而，f-tAFP并不广为人知，可能是因为它们的含量低于进化中不同的、对冰冻敏感的同类。因此，我们将使用现代技术，包括‘基因组’策略来克服这一挑战，以便从几个生物体中分离并随后鉴定新的f-tAFP。这些包括草以外的植物、昆虫和来自北极群落的细菌。*我们认为，这些物种包含可用于提高作物耐寒性的“配方”。这一点很重要，因为在过去几十年里，由于气候变化，冻融事件的频率有所增加。这些蛋白质还将用于动物组织的超低温保存。短期和长期目标将利用我们克隆的f-tAFP序列，以了解它们与冰和结晶气体水合物的相互作用。在未来，这些f-tAFP将在一些全球感兴趣的应用问题中找到用途。总体而言，我们的研究计划为实验和假设检验奠定了新的基础，这些调查鼓励不同的学生进行令人兴奋的发现计划。

Survival at low temperatures in some overwintering organisms such as certain fish, plants and insects, is enhanced by the expression of a number of genes including those for the unique antifreeze proteins (AFPs). AFPs bind to forming ice crystals and this patchy cover prevents ice growth. In freeze-susceptible species when temperatures drop further, AFPs can no longer ‘hold' ice crystals and explosive growth follows, leading to freezing and then death. In contrast, freeze-tolerant species freeze at higher sub-zero temperatures with AFPs keeping individual ice crystals small and offering cell protection. Ironically, considering the wealth of biochemical data on AFPs, the actual contribution of these proteins to freeze survival has not been established. We are now the first to do this by producing genetically-engineered brome grass with the AFPs ‘knocked down'. We must optimize this system in order to more fully explore the importance of these proteins. Our research suggests that AFPs from freeze-tolerant grasses (f-tAFPs) contribute to a complex web of low temperature, pathogen and immune interactions. We therefore postulate that f-tAFPs have multiple roles for: (i) protection of cell integrity by organizing ice, (ii) ‘spoiling' the nucleation of ice by bacteria, and (iii) a more general immune response against low-temperature tolerant bacteria. Innovatively, we expect that freeze-tolerant insects and microbial communities will show that other f-tAFPs also evolved as part of an ‘immune response'. However, f-tAFPs are not well known, likely because they are less abundant than their evolutionary-distinct freeze-susceptible counterparts. Thus we will use modern techniques including ‘omic' strategies to overcome this challenge in order to isolate, and subsequently characterize, novel f-tAFPs from several organisms. These include plants other than grasses, an insect and bacteria from an Arctic community.*******We submit that these species contain ‘recipes' that can be applied to increase freeze tolerance in crops. This is important since freeze-thaw events have increased in frequency over the last few decades due to climate change. These proteins will also have utility for the cryopreservation of animal tissues. Short and long-term goals will utilize our cloned f-tAFP sequences in order to understand their interactions with ice as well as crystalline gas hydrates. In the future, these f-tAFPs will find utility in a number of applied problems of global interest. Overall, our program of research establishes a new foundation for experimentation and hypothesis testing, with these enquiries encouraging diverse students to an exciting program of discovery.***