Functional studies of the stressosome

为了生存，所有的有机体都必须对环境的变化做出反应。所有生物体向自己发出信号表明需要对生长条件的变化做出反应的最重要的方式之一是通过蛋白质磷酸化来调节。在这里，通过磷酸化对调节蛋白进行共价修饰会改变目标蛋白的性质，从而改变其功能。磷酸化是由称为激酶的酶来执行的，它需要ATP作为磷酸化反应的辅助因子。枯草芽孢杆菌及其近缘细菌的应激反应也受到磷酸化的调节。对压力的反应为细胞提供了一种针对广泛的化学和物理侮辱的保护机制。最终，压力信号通路控制着分子机器--RNA聚合酶的活动，它是启动蛋白质合成的第一步所必需的，这些蛋白质共同作用，为细胞提供对施加的压力的抵抗力。向细胞表明其环境正在恶化的信息处理途径的关键是一个被称为“应激体”的大结构。应激体由几种蛋白质组成，它们一起将应激体组装成一个类似病毒的小结构。这种“迷你病毒”中的蛋白质协同作用，在压力前捕获一个关键的激酶，然后在压力下释放它，在最终激活RNA聚合酶的信号系统中发挥作用。我们目前正在确定各种功能状态下应激体的结构，这些正在进行的研究正在挑战我们回答这些结构提出的问题。这些答案只有通过结合使用经典生化方法和细菌细胞生物学技术进行功能研究才能得到。例如，我们首先需要确定应激体存在的原因--也许它的目的是对压力做出比压力信号大得多的反应，以确保细胞的健康得到维持。我们的目标是破坏应激体复合体的形成，以了解这种破坏对细菌细胞适当响应压力的能力的影响。此外，压力信号通路中还缺少一些成分，例如，单细胞生物体究竟是如何感知压力的？通过用一种蛋白质标记关键的信号转导激酶，我们可以找到关于应激受体的细胞位置和身份的线索，这种蛋白质将把激酶染成绿色，并使我们能够在细胞周期和对应激的反应期间使用显微镜追踪其位置。最后，在许多其他细菌中也发现了应激体蛋白，我们将证明它们确实在调节其他重要的细胞过程中发挥作用。

For survival, all organisms must respond to changes in their environment. One of the most important ways by which all organisms signal to themselves that a response is required to a change in growth conditions is mediated by protein phosphorylation. Here, the covalent modification of regulatory proteins by phosphorylation changes the properties of the target protein and hence its functions. Phosphorylation is performed by enzymes called kinases, which require ATP as a co-factor for the phosphorylation reaction. The stress response of the bacterium Bacillus subtilis and its close relatives is also regulated by phosphorylation. The response to stress provides the cell with a protective mechanism against a wide range of chemical and physical insults. Ultimately the stress signaling pathway controls the activity of the molecular machine, RNA polymerase, which is required to initiate the first step in the synthesis of the proteins that together act to provide the cell with its resistance against the imposed stress. Key to the information-processing pathway that indicates to the cell that its environment is deteriorating is a large structure called the 'stressosome'. The stressosome is composed of several proteins that together assembles the stressosome into a small virus-like structure. The proteins in this 'mini-virus' act in concert to trap a key kinase prior to stress and then to release it on stress to act in the signaling system that ultimately activates RNA polymerase. We are currently determining the structure of the stressosome in a variety of functional states and these on-going studies are challenging us to answer questions that the structures pose. These answers will only be obtained by functional studies using classical biochemical approaches in combination with bacterial cell biology techniques. For instance, we need to establish why the stressosome exists in the first place - perhaps its purpose is to provide a response to stress that is much greater in magnitude than the stress signal to guarantee that the health of the cell is maintained. We will aim to destroy the formation of the stressosome complex to see the impact on this disruption on the ability of the bacterial cell to respond appropriately to stress. Furthermore, there are missing components in the stress-signalling pathway, for instance how is stress actually perceived by a single-celled organism? Clues to the cellular location and identity of stress-receptors may be found by tagging the key signalling kinase with a protein that will colour the kinase green and enable us to track its location using a microscope during the cell cycle and during the response to stress. Finally, stressosome proteins are also found in many other bacteria and they are proposed to, and we will demonstrate that they do play roles regulating other important cellular processes.