Genes and their products for low temperature stress resistance

一些越冬生物如某些鱼类、植物和昆虫在低温下的存活率通过许多基因的表达而提高，包括那些独特的抗冻蛋白（AFP）。AFP与冰晶结合，这种片状覆盖物阻止了冰的生长。在易冻物种中，当温度进一步下降时，AFP不再能“保持”冰晶，随之而来的是爆炸性生长，导致冻结，然后死亡。相比之下，耐冷冻的物种在更高的零度以下温度下冷冻，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，这些蛋白质对冷冻存活的实际贡献尚未确定。我们现在是第一个通过生产基因工程雀麦草来做到这一点的。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，抗冻草（f-tAFPs）的AFP有助于低温，病原体和免疫相互作用的复杂网络。因此，我们假设f-tAFPs具有多种作用：（i）通过组织冰来保护细胞完整性，（ii）通过细菌“破坏”冰的成核，以及（iii）针对耐低温细菌的更普遍的免疫应答。创新性地，我们期望耐冷冻昆虫和微生物群落将表明其他f-tAFPs也作为“免疫反应”的一部分进化。然而，f-tAFPs并不为人所知，可能是因为它们比进化上不同的冷冻敏感对应物更不丰富。因此，我们将使用现代技术，包括“组学”策略，以克服这一挑战，以分离，并随后表征，新的f-tAFPs从几种生物体。这些包括除草以外的植物，一种昆虫和来自北极社区的细菌。我们认为，这些物种包含的“食谱”，可以应用于提高作物的抗冻性。这一点很重要，因为在过去几十年中，由于气候变化，冻融事件的频率增加了。这些蛋白质也可用于动物组织的冷冻保存。短期和长期目标将利用我们克隆的f-tAFP序列，以了解它们与冰以及结晶气体水合物的相互作用。在未来，这些f-tAFPs将在一些全球感兴趣的应用问题中找到效用。总的来说，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.***