The chaperone cycle of fibroblast growth factor receptor kinases in molecular detail

随着我们身体中的细胞老化或遇到外部压力，如来自饮食或环境的毒素，细胞内的蛋白质可能会累积损伤(例如突变)或表达水平的变化，导致它们形成聚集体，对细胞功能和生存造成损害。老年性疾病，如阿尔茨海默氏症和帕金森氏症，是与某些神经细胞组内形成大型聚集蛋白质结构有关的疾病的例子，导致表现为痴呆、震颤或失去运动控制。作为对压力的反应，细胞编组了一组称为伴侣的蛋白质，其功能是拯救这种异常折叠或聚集的蛋白质。然而，通常情况下，伴侣反应本身可能会通过细胞损伤的积累而受到损害，这可能会削弱有机体修复蛋白质聚集的能力，并导致随着年龄的增长而导致疾病的进展。伴侣反应的一个臂涉及HSP90蛋白，它被包括CDC37在内的各种辅助伴侣蛋白辅助识别“客户”蛋白。CDC37是参与细胞信号转导的重要蛋白家族的特异性辅伴蛋白，称为蛋白激酶。受体酪氨酸激酶(RTK)是一种从细胞外到细胞内传递信号的‘信号’激酶，包括本研究的主题成纤维细胞生长因子受体激酶(FGFRs)。FGFRs在胚胎发育、伤口愈合、新血管生成等过程中发挥着重要作用，其形态的改变导致了几种癌症的发生。我们感兴趣的是建立FGFRs如何与CDC37-HSP90伴侣系统相互作用的分子细节，特别是CDC37如何能够区分需要伴侣干预的FGFRs形式(例如，导致癌症等疾病的改变形式的FGFRs)和不需要伴侣干预的形式(例如大多数FGFRs的野生型变体)。我们的项目寻求通过使用一系列尖端技术和方法来填补我们知识中的一些重要空白。特别是，由于新的技术和方法，低温电子显微镜(CRYO-EM)最近在结构细节水平上经历了戏剧性的改进，在利兹和伦敦大学学院，我们可以获得最新一代的低温电子显微镜设备和专业知识。核磁共振和质谱学等其他结构技术在能力方面也取得了重大进展，我们在利兹和伦敦大学学院拥有专业知识和最先进的设备来承担这项工作。此外，在过去的5年里，我们已经建立了强大的试点数据体系，生成了所需的组件，特别是蛋白质复合体，这通常是这些具有挑战性的项目中的限制步骤。了解CDC37和带有FGFR激酶及其改变形式的伴侣系统在分子水平上的“结合规则”将有助于我们不仅了解伴侣系统在这一重要类别的蛋白质客户(RTK)方面的正常功能，而且还有助于我们了解它如何在急性或累积细胞应激中变得不堪重负或被颠覆。这样的理解也有助于有选择地针对伴侣系统的努力，以治疗由致癌激酶驱动的癌症，这些致癌蛋白依赖伴侣干预来维持其活性。

As cells in our body age or encounter external stresses, such as toxins derived from diet or the environment, proteins within the cell can accumulate damage (for example mutations) or alterations in expression levels that can cause them to form aggregates that are damaging to cell function and survival. Diseases of aging such as Alzheimer's and Parkinson's are examples of conditions associated with the formation of large, aggregated protein structures within certain groups of neuronal cells, leading to manifestations including dementia, tremor or loss of motor control. In response to stresses, cells marshal a set of proteins called chaperones whose function is to rescue such aberrantly folded or aggregated proteins. Often, however, the chaperone response itself can become compromised through accumulation of cellular damage, which can blunt the organism's ability to remediate protein aggregation and lead to disease progression with advancing age.One such arm of the chaperone response involves the protein HSP90, which is assisted in its recognition of 'client' proteins by various co-chaperones that include Cdc37. Cdc37 is the specific co-chaperone for an important family of proteins involved in cellular signalling known as kinases. Receptor tyrosine kinases (RTKs) are a type of 'signalling' kinase that transduce signals from outside to inside cells and include fibroblast growth factor receptor kinases (FGFRs), the subject of this study. FGFRs are important in embryonic development, wound-healing, generation of new blood vessels, and in altered forms are responsible for driving several types of cancer. We are interested in establishing the molecular details of how FGFRs interact with the Cdc37-HSP90 chaperone system, and in particular in how Cdc37 is able to distinguish between forms of FGFRs that require chaperone intervention (for example, altered forms of FGFRs responsible for diseases such as cancer) and those that do not (for example, wild-type variants of most FGFRs).Our project seeks to fill in some of the important gaps in our knowledge, by using a range of cutting-edge technologies and approaches. In particular, cryo-electron microscopy (cryo-EM) has recently undergone a dramatic improvement in the level of structural detail it can provide, thanks to new technologies and methods, and in Leeds and UCL we have access to the latest-generation cryo-EM equipment and expertise. Other structural techniques such as NMR and mass spectrometry have also seen significant advances in capability, and again, we have expertise and state-of-the-art equipment in Leeds and UCL to undertake this work. Moreover, over the last 5 years we have built up a strong body of pilot data, generating the components required, in-particular the protein complexes, which are often the limiting step in these challenging projects.Understanding the molecular-level 'rules of engagement' of Cdc37 and the chaperone system with FGFR kinases and their altered forms will help us to understand not only the normal functioning of the chaperone system in respect of this important class of protein clients (RTKs), but also how it might become overwhelmed or subverted in acute or cumulative cellular stress. Such understanding can also assist in efforts to target the chaperone system selectively to treat cancers driven by oncogenic kinases that are 'addicted' to chaperone intervention to maintain their activity.