Mechanism and Function of ISG15 Conjugation

描述（由申请人提供）：1型干扰素（IFN）应答是抵御病毒感染的第一道防线。在ifn刺激的细胞中，数百种蛋白质被诱导，这些蛋白质介导了广泛的抗病毒作用。ISG15是最早被鉴定的ifn诱导蛋白之一，也是第一个被发现的泛素样蛋白（ubitin -like protein, Ubl），但其生化功能及其抗病毒活性的基础多年来一直未被明确。ISG15对多种人类病毒具有抗病毒活性，包括流感病毒、逆转录病毒、信德病、埃博拉病毒，ISG15与其他蛋白质结合的能力可能是其在所有情况下的抗病毒活性所必需的。在ifn刺激的细胞中，ISG15与数百种细胞蛋白结合，而单个ifn诱导的连接酶Herc5介导几乎所有这些靶蛋白的结合。基于广泛的初步数据，我们假设Herc5是核糖体相关蛋白和ISGylates蛋白以共翻译的方式，几乎没有靶蛋白特异性。我们进一步假设，在干扰素反应的背景下，新翻译的病毒蛋白而不是细胞蛋白是该系统的主要目标，并且isg酰化是一种灭活病毒蛋白功能的尝试。这一提议的目的将检验这两种假设。干扰素反应是一个多方面的防御屏障，可以保护细胞免受各种感染因子的侵害，但只有少数干扰素诱导的蛋白质在分子细节上得到了表征。此外，干扰素用于治疗病毒感染（如乙型肝炎和丙型肝炎以及乳头瘤病毒），以及某些类型的癌症和多发性硬化症，但尚不清楚哪些干扰素诱导的蛋白质介导有用的治疗效果，哪些介导这些疗法的臭名昭着的副作用。进一步了解ISG15偶联的机制和功能将为在这些临床环境中上调或下调isg酰化活性提供机会。

DESCRIPTION (provided by applicant): The type 1-interferon (IFN) response is the first line of defense against viral infections. Hundreds of proteins are induced in IFN-stimulated cells, and these proteins mediate a wide spectrum of anti-viral effects. ISG15 was one of the first IFN-induced proteins to be identified and the first ubiquitin-like protein (Ubl) to be discovered, however its biochemical function and the basis of its antiviral activities have remained largely uncharacterized for many years. ISG15 has anti-viral activity against a wide range of human viruses, including influenza, retroviruses, sindbis, ebola, and the ability of ISG15 to be conjugated to other proteins is likely to be essential for its anti-viral activity in all cases. ISG15 is conjugated to hundreds of cellular proteins in IFN-stimulated cells, and a single IFN-induced ligase, Herc5, mediates conjugation to nearly all of these target proteins. Based on extensive preliminary data, we hypothesize that Herc5 is ribosome-associated and ISGylates proteins in a co-translational manner, with little target protein specificity. We further hypothesize that in the context of an interferon response, newly translated viral proteins, rather than cellular proteins are the primary targets of this system, and that ISGylation is an attempt to inactivate the function of viral proteins. The aims of this proposal will test both of these hypotheses. The interferon response is a mutli-faceted defensive shield that protects cells against a wide range of infectious agents, yet only a small number of IFN-induced proteins have been characterized in molecular detail. In addition, interferon is used therapeutically in treating viral infections (e.g., hepatitis B and C and papillomaviruses), as well as certain types of cancers and multiple sclerosis, yet it is not known which IFN-induced proteins mediate useful therapeutic effects and which mediate the notorious side effects of these therapies. A further understanding of the mechanism and function of ISG15 conjugation will present opportunities for either up- or down-modulating ISGylation activity in these clinical settings. PUBLIC HEALTH RELEVANCE: ISG15 conjugation is now recognized to be an important aspect of the innate immune response against a wide range of human viral infections, and interferon therapies are approved for many disease states. It is essential to characterize the many facets of the interferon response system, including the ISG15 pathway, in order to improve and modulate anti-viral and interferon therapies.