Functions and Mechanisms of Deubiquitinating Enzymes

描述（由申请人提供）：真核生物具有高度保守的酶系统，用于将泛素（Ub）连接到蛋白质上。此外，与Ub不同但与Ub相关的多肽，称为Ub样蛋白或Ubls，也可以附着在蛋白质上。连接到每个Ubl具有独特的机制和功能结果。SUMO （small Ub-related modifier）是一种高度分化的Ubl， SUMO连接系统在许多生物中起着至关重要的作用，包括对人类生物学的重要贡献。Ub和SUMO对蛋白质的附着在体内都可以迅速逆转，并且专门的蛋白酶负责这些裂解反应。PI一直在分析脱泛素酶（DUB）家族，主要存在于酵母酿酒酵母中，并且在PI的实验室中发现了第一个sumo特异性蛋白酶ulp。ULP类蛋白酶是小鼠胚胎发生所必需的，并且在几种人类癌症中过度表达。因此，这些酶已成为药物开发的有吸引力的靶点。

DESCRIPTION (provided by applicant): Eukaryotes 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 attached to proteins. Ligation to each Ubl has unique mechanistic and functional consequences. SUMO (small Ub-related modifier) is a highly divergent Ubl, and the SUMO ligation system has crucial roles in many organisms, including important contributions to human biology. Both Ub and SUMO attachment to proteins can be rapidly reversed in vivo, and specialized proteases are responsible for these cleavage reactions. The PI has been analyzing the deubiquitinating enzyme (DUB) family, primarily in the yeast Saccharomyces cerevisiae, and the first SUMO-specific proteases, the ULPs, were discovered in the PI's laboratory. ULP- class proteases are essential for embryogenesis in the mouse and are overexpressed in several human cancers. Therefore these enzymes have emerged as attractive targets for drug development. The long-range objective of the project is to gain a molecular understanding of the physiological and mechanistic roles played by DUBs and ULPs in vivo. In this renewal application, the proposed experiments are concentrated on SUMO modification in yeast and on the contributions of the two yeast desumoylating enzymes, Ulp1 and Ulp2, to SUMO system function. Mutation of either ULP has strong effects on growth and division, and Ulp1, like SUMO itself, is essential for cell-cycle progression. In broad terms, the goals are two-fold: Determine the molecular basis for key regulatory functions of the Ulp1 and Ulp2 enzymes and elucidate the molecular features of these SUMO proteases that are responsible for their dramatic differences in specificity and activity. Recent data on Ulp1 and Ulp2 have directed the studies into several specific areas of biological regulation. Based on these new findings, the following Aims are proposed: (1) Examine the function of Ulp1 at the nuclear pore complex, particularly its role in pre-mRNA nuclear retention; (2) Determine novel regulatory features of Ulp2, especially its role in chromatin regulation, and determine the contributions of Ulp2 noncatalytic domains to its in vivo regulation; and (3) Examine how SUMO attachment to substrates apparently promotes their ubiquitination by the heterodimeric Hex3-Slx8 Ub ligase. PUBLIC HEALTH RELEVANCE: The growth and behavior of human cells, like those of virtually all complex organisms, is controlled by rapid attachment and removal of small specialized proteins (called ubiquitin-like proteins) to and from other proteins. Defects in the enzymes that control these processes are known to cause human developmental abnormalities, neurodegenerative disorders, and many different forms of cancer. This project aims to deepen our understanding of the enzymes that detach certain ubiquitin-like proteins from their partners, with the long-term goal of developing therapies to treat patients suffering from cancer and other diseases.