Progesterone Receptor and the hsp90 Chaperone Pathway

孕激素受体和HSP90分子伴侣途径。与大多数类固醇受体和其他几种细胞信号蛋白一样，禽孕酮受体(PR)在胞浆提取物中处于非活性状态时，与热休克蛋白HSP90复合。这种复合体不是通过简单的结合形成的，而是通过一个需要ATP水解和几个额外蛋白质的过程形成的。其他参与受体复合体形成的蛋白质包括HSP70、三个HSP70辅助伴侣、Hsp40、Hip和Hop、四个HSP90结合的TPR蛋白和p23。我们已经开发了一个化学定义的系统，使用纯化的蛋白质，它能够与HSP90组装PR复合体。这个体外组装过程只需要上述五种蛋白质，这一过程似乎要经过三个步骤才能产生具有激素结合活性的PR。这些研究的目的是为了加深我们对受体复合体的组装、它们在激素结合时的解离以及受体相关蛋白的功能意义的理解。这将通过广泛使用体外定义的组装系统来实现。PR复合体形成的每个步骤的事件将根据蛋白质相互作用的动力学和对ATP的利用来描述。将对系统进行修改，以允许研究激素依赖的PR从HSP90中解离。这可能需要确定解离过程所需的新因素。最后，我们将确定PR的结构域或元件是与HSP70和HSP90结合的相互作用部位，并研究促进热休克蛋白识别的特性。这些研究将导致类固醇受体激活过程的机制描述，并为理解分子伴侣HSP70和HSP90的作用机制提供有价值的信息。

Progesterone receptor and the hsp90 chaperone pathway. When in its inactive form in cytosol extracts, the avian progesterone receptor (PR), like most steroid receptors and several other cell signaling proteins, is complexed with the heat shock protein hsp90. This complex does not form through a simple association, but through a process that requires ATP hydrolysis and several additional proteins. Other proteins involved in receptor complex formation are hsp70, three hsp70 co-chaperones, hsp40, Hip and Hop, four hsp90-binding TPR proteins and p23. We have developed a chemically defined system with purified proteins that has the capacity to assemble PR complexes with hsp90. Only five of the above proteins are needed for this in vitro assembly process which appears to proceed through three steps to generate PR with hormone binding activity. The objective of the proposed studies is to further our understanding on the assembly of receptor complexes, their dissociation upon hormone binding, and the functional significance of the receptor-associated proteins. This will be accomplished by extensive use of the in vitro defined assembly system. Events at each step of PR complex formation will be characterized in terms of the dynamics of protein interactions and the utilization of ATP. Modifications in the system will be made that allow the study of hormone- dependent dissociation of PR from hsp90. This will probably require the identification of new factors needed for the dissociation process. Finally, we will characterize the domains or elements of the PR that are interaction sites for binding hsp70 and hsp90 and investigate the properties which promote heat shock protein recognition. These studies should lead toward a mechanistic description of the steroid receptor activation process and should provide valuable information for understanding the mechanisms of action of the molecular chaperones hsp70 and hsp90.