Relating protein interaction networks to physiology by systematic mutant analyses

HSP90是真核生物中一种重要的伴侣蛋白，有助于产生和维持活性状态。 不成比例地链接到信令过程的一组选定的底物或客户端 包括许多促进癌症和真菌中出现抗药性的因素。为了这些 原因，操纵Hsp90伴侣系统的抑制策略有望带来许多潜力 治疗方面的好处。靶向Hsp90 ATPase位点的抑制剂有效地限制了 在真菌中出现耐药性，并显示出作为抗癌药物的前景。 然而，ATPase抑制剂阻断了Hsp90的所有已知功能，导致了不良的副作用 效果。在我们之前的工作中，我们在以下位置生成了所有可能的突变的综合文库 Hsp90中709个位置的每一个，并量化每个变体对酵母生长的影响 Rate是所有客户蛋白质上整合的整体网络功能的读数。我们将在此基础上 本工作旨在研究Hsp90与客户端的交互机制。我们将确定四个具体的 客户受到Hsp90突变的影响，这将识别Hsp90上可能 目标是抑制特定的客户端。我们将对特定客户进行深度突变扫描 确定决定Hsp90依赖的生物物理特征的蛋白质。此外，我们 已经开发出一种新的方法来量化Hsp90突变的显性影响，这 我们正在利用它作为一条强大的新途径来理解它的作用机制。我们的研究将 提供对蛋白质-蛋白质相互作用网络的重要机制洞察 在生物学和医学上都很重要。

Hsp90 is an essential eukaryotic chaperone that helps to produce and maintain the active state of a select set of substrates or clients that are disproportionately linked to signaling processes including many that promote cancer and the emergence of drug resistance in fungi. For these reasons, inhibitor strategies to manipulate the Hsp90 chaperone system promise many potential therapeutic benefits. Inhibitors targeting the ATPase site of Hsp90 effectively limit the emergence of drug resistance in fungi and show promise as anti-cancer agents. However, ATPase inhibitors block all known functions of Hsp90, leading to undesirable side effects. In our previous work, we generated a comprehensive library of all possible mutations at each of the 709 positions in Hsp90 and quantified the effects of each variant on yeast growth rate as a readout of overall network function integrated over all client proteins. We will build on this work to investigate Hsp90-client interaction mechanism. We will determine how four specific clients are impacted by Hsp90 mutations, which will identify sites on Hsp90 that could be targeted to inhibit specific clients. We will perform deep mutational scanning on specific client proteins to identify the biophysical features that determine Hsp90 dependence. In addition, we have developed a novel approach to quantify the dominant effects of Hsp90 mutations, which we are using as a powerful new route to understand its mechanism of action. Our studies will provide important mechanistic insights into a protein-protein interaction network that is biologically and medically important.