Functions and Mechanisms of Deubiquitinating Enzymes

真核细胞具有高度保守的泛素连接酶系统 (Ub)到蛋白质。此外，与Ub不同但相关的多肽，称为Ub样 蛋白质或Ubls也可以与蛋白质缀合。与每个Ubl的连接具有独特的 机械性和功能性后果。相扑是一种高度分化的Ubl。相扑 和Ub在许多生物体中起着至关重要的作用， 人类生物学这些修饰被专门的蛋白酶逆转。下 在这项资助的支持下，我们的研究小组一直在分析两种去泛素化酶 （DUBs）和一组对SUMO特异的Ubl特异性蛋白酶（ULP）。蛋白质我们 从酵母到人类都是保守的。的长期目标 该项目旨在从分子水平上了解 由DUBs和ULP播放。 在这个更新应用程序中，所提出的实验集中在两个不同的主题。 第一个是实验室最近通过演示进入的一个令人兴奋的新领域 来自专性细胞内细菌Wolbachia pipientis的特定DUB， 负责一种长期神秘的昆虫生殖寄生类型， 细胞质不亲和性（CI）。DUB作为一种毒素发挥作用，编码为两个- 基因Wolbachia操纵子。这种新的联系对以下方面具有潜在的重要意义： 控制害虫和疾病媒介。第二个主题集中在如何 一种神秘的酵母SUMO蛋白酶Ulp 2，对SUMO系统的功能有贡献 在转录中以及它的丢失如何意外地导致特定的多染色体 非整倍性。后者可以使我们理解的基本功能（S） Ulp 2在细胞分裂中的作用，以及对人类细胞非整倍性的了解，这是细胞分裂的标志。 癌建议的具体目标如下： (1)研究CidB（Wolbachia Ulp 1/C48家族DUB）诱导CI的机制， 使用果蝇和酵母模型。(2)确定沃尔巴克氏体CinB蛋白， 一种假定的核酸酶，也有助于CI，如果是这样，它的细胞分裂机制如何 破坏不同于CidB诱导的CI。(3)研究Ulp 2 SUMO蛋白酶 通过基因特异性和基因组方法调节酵母中的转录， 确定Ulp 2缺失诱导的特异性非整倍体的基础。

Eukaryotic cells have a highly conserved enzymatic system for the ligation of ubiquitin (Ub) to proteins. Moreover, polypeptides distinct from but related to Ub, called Ub-like proteins or Ubls, can also be conjugated to proteins. Ligation to each Ubl has unique mechanistic and functional consequences. SUMO is a highly divergent Ubl. Both SUMO and Ub have crucial roles in many organisms, including important contributions to human biology. These modifications are reversed by specialized proteases. Under the auspices of this grant, our group has been analyzing both deubiquitylating enzymes (DUBs), and a set of Ubl-specific proteases (ULPs) specific for SUMO. The proteins we have been studying are conserved from yeast to humans. The long-term objective of the project is to gain a molecular understanding of the physiological and mechanistic roles played by DUBs and ULPs. In this renewal application, the proposed experiments concentrate on two distinct topics. The first is an exciting new area recently entered by the lab through the demonstration that a specific DUB from an obligate intracellular bacterium, Wolbachia pipientis, is responsible for a long mysterious type of reproductive parasitism in insects called cytoplasmic incompatibility (CI). The DUB functions as a toxin encoded as part of a two- gene Wolbachia operon. This novel connection has potentially important implications for controlling insect pests and disease vectors. The second topic focuses on how an enigmatic yeast SUMO protease, Ulp2, contributes to the function of the SUMO system in transcription and how its loss, unexpectedly, leads to a specific multi-chromosome aneuploidy. The latter could lead us to an understanding of the essential function(s) of Ulp2 in cell division and insights into aneuploidy in human cells, which is a hallmark of cancer. The following Specific Aims are proposed: (1) Examine the mechanism of CI induced by CidB, a Wolbachia Ulp1/C48-family DUB, using both Drosophila and yeast models. (2) Determine if the Wolbachia CinB protein, a putative nuclease, also contributes to CI and if so, how its mechanism of cell division disruption differs from CI induced by CidB. (3) Investigate how the Ulp2 SUMO protease regulates transcription in yeast by gene-specific and genomic approaches, and determine the basis of the specific aneuploidy induced by loss of Ulp2.