Is cytokine signalling kinetically controlled by the dissociation rate of STAT dimers?

在动物和人类中，一个复杂的细胞间通讯系统协调器官和组织中不同细胞类型的活动。通信可以是通过直接细胞与细胞接触的短距离通信，也可以是需要不同化学性质的信号分子的长距离通信，这些信号分子从一个细胞释放并在生物体中循环，直到它们遇到它们的靶细胞。靶细胞可以捕获信号分子并启动生化反应，以信号分子指定的方式触发反应。我们关注的是由称为细胞因子的信号分子传递的信号，细胞因子是一组约40种不同的小蛋白质，通过调节基因的活性来影响其靶细胞。因此，响应于这些信号分子，某些基因被打开，而其他基因变得不那么活跃。细胞因子诱导的基因活性变化调节许多细胞类型的生长和分化，并保护它们免受病毒攻击。然而，细胞因子并不直接调节基因，而是通过靶细胞中称为STAT蛋白的中间体。因此，对丝氨酸应答基因的控制最终需要STAT蛋白的调节。不幸的是，细胞的常见故障和严重人类疾病如炎症或癌症的原因是过量的精氨酸诱导的信号传导。这通常是由于未能终止其他正常信号。因此，研究STAT蛋白的活性如何被关闭是当前基础和医学研究的重要课题。为了控制基因活性，STAT需要与编码相应基因的DNA片段结合。值得注意的是，STAT并不作为单个分子结合DNA，而是两个STAT分子需要组装成对，然后才能结合和调节基因。因此，成对STAT分子的断裂是细胞因子信号下调的关键事件。在我们最近对STAT自组装的研究中，我们发现成对的STAT 1分子非常稳定，而且寿命出乎意料地长。特别有趣的是，成对的STAT 1分子的断裂和细胞因子信号的下调是偶联的，并且似乎以非常相似的速率进展。如果情况确实如此，STAT对的分离将是限制细胞终止信号流的能力的决定性步骤。药理学干预的潜力是显而易见的;任何加速成对STAT分子分离的措施都将对抗过度信号传导，具有非常理想的治疗效果。然而，这方面的蜂窝信号处理已经收到很少的关注日期。在这里，我们因此建议探索细胞因子信号的终止和配对STAT分子的断裂之间的联系。因此，我们将首次确定STAT配对和非配对的比率。这些试管分析将用两种不同的STAT进行，我们以前的研究已经证明了它们具有显著不同的自组装。然后，我们将使用另一组实验来分析STAT蛋白在活细胞中的天然环境中的行为;并将这些数据与我们用分离的蛋白质获得的结果进行比较。这将揭示解配对是否仅由STAT的固有分子特性决定，或者是否存在调节该过程的活细胞中的机制。最后，我们将试图确定DNA是否以及如何影响配对STAT 1的形状，以更好地了解基因结合如何影响STAT分子的分离。总的来说，这些实验将揭示细胞通讯的基本原理，并可能建立一个新的调节机制，在细胞信号转导的STAT的un-pairing。这可以启动开发利用细胞信号处理的全新方面的药物。

In animals and humans, an intricate cell-to-cell communication system coordinates the activities of the different cell types in organs and tissues. Communication can be short range by direct cell-to-cell contact, or over longer distances that require signalling molecules of diverse chemical natures, which are released from one cell and circulate through the organism until they come across their target cells. The target cells can capture the signalling molecules and initiate biochemical reactions that trigger a response in a manner specified by the signalling molecule. We are concerned with signals that are conveyed by signalling molecules called cytokines, a group of about 40 different small proteins that affect their target cells by modulating the activity of genes. Thus, in response to these signalling molecules certain genes are switched on while others become less active. Cytokine-induced changes in gene activity regulate the growth and differentiation of many cell types and protect them against viral attack. However, the cytokines do not regulate genes directly, but through intermediaries in the target cells named STAT proteins. Therefore, control of cytokine-responsive genes ultimately requires the regulation of STAT proteins. Unfortunately, a common malfunction of cells and cause of serious human diseases such as inflammation or cancer is excessive cytokine-induced signalling. Often this is due to failure to terminate otherwise normal signals. For this reason the study of how the activity of STAT proteins is turned off is an important topic of current basic and medical research. In order to control gene activity the STATs need to bind to those segments of DNA that encode the respective gene. Notably, the STATs do not bind DNA as single molecules, but two STAT molecules need to assemble into pairs, which are then competent to bind and regulate genes. Therefore the breaking-up of paired STAT molecules is the crucial event in the down-regulation of cytokine signals. In our recent study of STAT self-assembly we made the discovery that pairs of STAT1 molecules are remarkably stable and unexpectedly long-lived. It is particularly intriguing that the breaking-up of paired STAT1 molecules and the down regulation of cytokine signals are coupled and appear to progress at very similar rates. If this was indeed the case, the separation of STAT pairs would be the decisive step that limits the cell's capability to terminate the signal flow. The potential for pharmacological intervention is obvious; any measure to accelerate the separation of paired STAT molecules would counter excessive signalling-with highly desirable therapeutical effects. However, this aspect of cellular signal processing has received little attention to date. Here, we thus propose to explore the link between the termination of cytokine signals and the breaking-up of paired STAT molecules. We are therefore going to determine for the first time the rates of STAT pairing and un-pairing. These test-tube analyses will be done with two different STATs for which our previous study has demonstrated significantly different self-assembly. We will then use another set of experiments to analyse the behaviour of STAT proteins in their native environment in living cells; and compare those data with our results obtained with the isolated proteins. This will reveal whether un-pairing is determined solely by the inherent molecular properties of STATs, or if mechanisms exist in living cells that modulate this process. Finally, we will try to determine whether and how DNA affects the shape of paired STAT1 to better understand how gene binding influences the separation of STAT molecules. Collectively, these experiments will reveal basic principles of cellular communication and may establish un-pairing of STATs as a novel regulatory mechanism in cell signalling. This can initiate the development of drugs that exploit an entirely novel facet of cellular signal processing.