Roles and regulation of aqua/glyceroporins in a freeze tolerant amphibian

冷冻保存生物组织在从生殖技术到濒危物种保护等领域都有应用。然而，尽管科学家们可以成功地低温保存像卵子和胚胎这样的小型生命结构，但将这些策略应用于更大的结构仍然具有挑战性。自然界可以为克服这些挑战提供见解。许多“冷血”动物在自然环境中会遇到低于冰点的温度，在脊椎动物中，一些物种已经进化出了生理和生化策略来在寒冷中生存。这个项目将研究其中一种物种，Cope？灰树蛙。冰冻给动物带来了令人生畏的挑战，包括冰对身体的潜在伤害，血液冻结和解冻时组织氧合的波动，以及液态水冻结和体液浓度变化对分子和细胞结构的破坏。生存这些挑战的关键是调节水和溶解溶质的分布。冰的形成必须限制在细胞外的液体中，因此当冻结开始时，水必须能够迅速离开细胞。此外，一些溶质，如甘油具有低温保护作用，有助于在冷冻过程中保持分子和生理的完整性。灰色树蛙为揭示这些能力背后的机制提供了一个自然的模型。在凉爽的秋季，它们会积累甘油。它们还调节促进水和甘油穿过细胞膜的蛋白质（“水通道蛋白”）的表达。该项目将评估这些分子机制是如何被调节的，对细胞特性的影响是什么，以及这如何有助于整个动物的抗冻性。这种综合方法为培养从高中到博士水平的学生分子生理调控的方法和思想提供了一个很好的框架，它将对保持组织冷冻存活的挑战产生新的见解。

Preservation of biological tissues by freezing has application in fields ranging from reproductive technology to conservation of endangered species. However, while scientists can successfully cryopreserve small living structures like eggs and embryos, application of those strategies to larger structures remains challenging. The natural world can provide insights into overcoming those challenges. Many "cold-blooded" animals encounter sub-freezing temperatures in their natural environment and, among the vertebrates, a few species have evolved physiological and biochemical strategies to survive freezing. This project will study one of those species, Cope?s gray treefrog. Freezing imposes daunting challenges for an animal, including potential physical damage from ice, fluctuating tissue oxygenation as the blood freezes and thaws, and disruption of molecular and cellular structures as liquid water freezes and body fluid concentrations change. A key to surviving these challenges is regulation of the distribution of water and dissolved solutes. Ice formation must be restricted to fluids outside of cells, and so water must be able to leave cells rapidly when freezing begins. In addition, some solutes, such as glycerol are cryoprotective and help to preserve molecular and physiological integrity during freezing. Gray treefrogs present a natural model for unraveling the mechanisms behind these capabilities. During the cooling autumn months they accumulate glycerol. They also regulate the expression of proteins ("aquaporins") that facilitate movement of water and glycerol across cell membranes. This project will evaluate how these molecular mechanisms are regulated, what is the resulting impact on cellular properties, and how this contributes to freeze tolerance of the whole animal. This integrative approach provides an excellent framework for training students from high school through PhD level in methods and ideas of molecular physiological regulation, and it will yield novel insights into the challenge of keeping tissues frozen alive.