Protein quality control at eukaryotic membranes.

内质网相关降解(ERAD)是ER蛋白被降解的过程 因为它们被错误折叠，或者因为它们的降解是生理调节的。的标志性底物 这个过程就是HMG-CoA还原酶，这是一种催化胆固醇合成限速步骤的酶。在……里面 除了降解代谢调节或错误折叠的蛋白质外，ERAD还起到应激反应系统的作用 缓解内质网压力。虽然ERAD的正常功能是靶向错误折叠或调节的蛋白质，但ERAD也是 被细菌和病毒病原体利用以获取细胞质。在正常的ERAD期间，蛋白质底物 在内质网管腔内被选择，穿过细胞膜移动到胞浆(逆转位)，在那里它们 是多泛素化的，从膜上提取并被蛋白酶体降解。从生理上讲- 在ERAD的控制下，选择调节底物进行降解和特定的细胞通路 是模棱两可的。最近的研究表明，一种名为Hrd1的中央保守的泛素连接酶形成一种 泛素门控蛋白传导通道，足以使错误折叠的管腔逆转移位 和完整的膜蛋白。自动素化门控机制是一个难题；在正常情况下 在这种情况下，泛素化会导致修饰蛋白的降解。然而，Hrd1相对稳定 这意味着存在不明机制来保护自动激活的Hrd1不被降解，并 逆转泛素门控激活。除了对Hrd1的调控外，许多相关的过程包括 ERAD的底物选择是神秘的。该提案的目标是1)定义ERAD系统如何 2)确定ERAD系统如何识别其目标，3)定义目标导向的特征 4)了解膜如何对ERAD功能起作用；5)确定 其他(非ERAD)整体膜蛋白质量控制系统的细胞功能。我们将使用 结合生物化学、细胞生物学、遗传学和蛋白质组学的多方面方法来解决这些中心问题 膜相关蛋白质量控制中的问题。我们将利用我们独特的体内和体外测试 (并继续设计创新的分析方法)来剖析这些真核系统的基本机制。一个 对系统如何运作(包括ERAD)及其监管过程的机械性理解 在蛋白质错误折叠的医学相关途径中建立这些系统作为可行的治疗靶点， 蛋白质失调和病原体劫持。

Endoplasmic reticulum-associated degradation (ERAD) is a process in which ER proteins are degraded, either because they are misfolded or because their degradation is physiologically regulated. The hallmark substrate of this process is HMG-CoA reductase, an enzyme catalyzing the rate-limiting step in cholesterol synthesis. In addition to degrading metabolically-regulated or misfolded proteins, ERAD functions as stress-response system to alleviate ER stress. While its normal function is to target misfolded or regulated proteins, ERAD is also exploited by bacterial and viral pathogens to gain access to the cytosol. During normal ERAD, protein substrates are selected within the ER lumen, moved across the membrane to the cytosol (retrotranslocation), where they are polyubiquitinated, extracted from the membrane and degraded by the proteasome. How physiologically- regulated substrates are selected for degradation and the specific cellular pathways under the control of ERAD are ambiguous. Recent work has demonstrated that a central, conserved ubiquitin ligase called Hrd1, forms a ubiquitin-gated protein-conducting channel that that is sufficient to allow retrotranslocation of misfolded lumenal and integral membrane proteins. The autoubiquitination-gating mechanism presents a conundrum; under normal circumstances, ubiquitination will result in degradation of the modified protein. However, Hrd1 is relatively stable meaning there are unidentified mechanisms in place to protect autoubiquitinated Hrd1 from degradation and to reverse the ubiquitin-gated activation. Along with the regulation of Hrd1, many associated processes including substrate selection by ERAD are mysterious. The goals of this proposal are to 1) define how the ERAD system is regulated, 2) determine how the ERAD system recognizes its targets, 3) define the features of targets directing them to ERAD (the degrons), 4) understand how the membrane contributes to ERAD function, and 5) identify the cellular function of other (non-ERAD) integral membrane protein quality control systems. We will use a multifaceted approach with biochemistry, cell biology, genetics, and proteomics to address these central questions in membrane-associated protein quality control. We will leverage our unique in vivo and in vitro assays (and continue to design innovative assays) to dissect the basic mechanisms of these eukaryotic systems. A mechanistic understanding of how the systems function (including ERAD) and the processes they regulate will establish these systems as viable therapeutic targets in medically relevant pathways of protein misfolding, protein misregulation, and pathogen hijacking.