Elucidating the transcriptional and post-translational systems that regulate snake venom assembly and that control autolysis and self-toxicity

毒蛇毒液由自然界最具组织破坏性的蛋白质组成。这些酶会导致被毒死的啮齿动物猎物的心血管迅速而灾难性的崩溃，在人类身上，会导致危及生命的出血和非凝血。然而，尽管这种致命的鸡尾酒毒素储存在蛇的毒腺中，但它受到某种程度的控制，因此蛇自己的组织不会受到影响。这种不同寻常的生物现象知之甚少，但显然是毒液作为毒蛇的一种捕食和防御/攻击性生存策略成功进化发展的核心。毒蛇蛋白毒性的控制研究很少，因为合适的基因和蛋白质鉴定技术直到最近才出现。以前，我们和其他人已经使用基本的生化技术证明，一组蛇毒酶保持静止状态，直到在同样未知的时间框架内被其他未知的蛇毒酶激活。人们还认为，毒液中含有三种关键的毒蛇毒素的抑制剂，但这一点还有待实验验证。我们最近在应用新的高通量方法方面开发了专业知识和资源，以识别新的蛇毒基因。在这里，在这类研究的第一次，我们的目标是提供第一个整体观点，即毒素和毒素调节毒腺基因是如何在毒液合成过程中协调的。在新的基因和蛋白质定量技术的辅助下，我们将首先鉴定构成模型蛇毒的基因及其蛋白质产物。接下来，我们将在几个时间间隔内监测这些基因和蛋白质水平的变化，时间跨度从蛇排出毒液到它重新合成完全合格的新毒液武器库的时间。使用计算机技术，我们将区分毒素和预测用于调节其毒性活动的蛋白质。我们将使用分子技术来确定控制(I)毒素和(Ii)毒素调节蛋白产生的遗传机制是否协调一致，以确保调节蛋白在各自的毒素之前产生。这项研究的结果将极大地提高我们对毒液组装和毒力调节的动力学和基本生物学原理的理解。

Viper venom consists of a mixture of nature's most tissue-destructive proteins. These enzymes cause rapid and catastrophic cardiovascular collapse in envenomed rodent prey and, in humans, life-threatening bleeding and non-clotting blood. Yet while this lethal toxin cocktail is stored in the snake's venom gland it is somehow controlled so that the snakes' own tissues are unaffected. This extraordinary biological phenomenon is very poorly understood but is clearly central to the successful evolutionary development of venom as a predation and defensive/offensive survival strategy for venomous snakes. The control of viper protein toxicity has been very little studied because the appropriate gene and protein identification technologies have only recently become available. Previously, using basic biochemical techniques we and others have shown that one group of venom enzymes are kept in a quiescent state until activated by other, unknown, venom enzymes in an equally unknown time frame. It is also believed venom contains inhibitors to three key viper venom toxins but this has yet to be experimentally verified. We have recently developed expertise and resources in the application of new high throughput methods to identify new viper venom genes. Here, in the first study of its kind, we aim to provide the first holistic view of how sets of toxin and toxin-regulating venom gland genes are orchestrated during venom synthesis. Assisted by new gene and protein quantification technologies, we will first identify the genes and their protein products that constitute a model viper venom. Next, we will monitor the changes in the levels of these genes and proteins at several intervals spanning the time from when the snake ejects venom to the time it re-synthesises a fully competent arsenal of new venom. Using computational techniques, we will distinguish between toxins and proteins predicted to regulate their toxic activity. We will use molecular techniques to identify whether the genetic machinery controlling the production of (i) the toxins and (ii) the toxin-regulatory proteins, are co-ordinated to ensure that the regulatory proteins are produced before the respective toxin. The results of this study will greatly improve our understanding of the dynamics and the fundamental biological principles regulating venom assembly and venom toxicity.