Smt3-conjugation in cell biology and development

细胞生物学和发育中的 Smt3 缀合

• 批准号: 6796751
• 负责人: ALBERT J COUREY
• 金额: $ 18.88万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-15 至 2006-12-10
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6796751
• 关键词: Drosophilidae; apoptosis; arthropod genetics; cell biology; chromosome movement; gene induction /repression; gene mutation; growth /development; lysine; mass spectrometry; protein protein interaction; protein sequence; site directed mutagenesis; tissue /cell culture; transcription factor; ubiquitin

DESCRIPTION (provided by applicant): Ubiquitin-like proteins are receiving increasing attention due to their recently discovered roles in diverse biological processes. One such protein, named Smt3 (also known as Sumo or Sentrin), has a wide range of suspected functions, including functions in transcriptional regulation, nuclear transport, and cytokinesis. Like ubiquitin, Smt3 becomes covalently attached to target proteins via isopeptide linkages to lysine residues. These targets include transcription factors, components of the nuclear pore complex, and components of the septin complex. While many Smt3-conjugation targets have been identified, studies of Smt3 function in metazoans has largely been limited to cell culture analysis. This proposal describes a combined biochemical, cell biological, and genetic analysis of Smt3-conjugation in Drosophila, with the goal being to determine the functional roles of this process in the developing organism. The specific aims are to: (1) Identify targets of Smt3-conjugation in Drosophila by expressing tagged forms of Smt3 in vivo and then using the tags to purify Smt3-conjugates from protein extracts; (2) Determine the results of perturbing Smt3-conjugation in cultured Drosophila cells on such processes as protein localization and transcriptional regulation; (3) Determine the developmental roles of Smt3-conjugation in the intact organism by using genetic and reverse genetic approaches to perturb Smt3-conjugation in vivo, with the goal of making direct links between observed phenotypes and specific targets of Smt3-conjugation. These studies have many implications for human health. For example, one recently discovered target of Smt3-conjugation is the promyelocytic leukemia protein PML. The conjugation of this protein to Smt3 appears to control its localization to discrete nuclear foci termed PML bodies. Recent experiments suggest that Drosophila nuclei may contain similar Smt3-dependent foci.

描述（由申请人提供）：泛素样蛋白正在接受 由于最近发现它们在不同领域中的作用而受到越来越多的关注 生物过程。其中一种蛋白质名为 Smt3（也称为 Sumo 或 Sentrin），具有广泛的可疑功能，包括以下功能： 转录调控、核运输和胞质分裂。就像泛素一样， Smt3 通过异肽键共价连接至靶蛋白 赖氨酸残基。这些靶标包括转录因子、 核孔复合体和隔膜复合体的成分。虽然很多 Smt3 缀合靶点已确定，Smt3 功能研究 后生动物很大程度上仅限于细胞培养分析。这个提议 描述了生物化学、细胞生物学和遗传分析的结合 果蝇中的 Smt3 缀合，目标是确定功能 这个过程在发育中的有机体中的作用。 具体目标是： (1) 确定 Smt3 缀合的靶标 果蝇体内表达标记形式的 Smt3，然后使用标记 从蛋白质提取物中纯化 Smt3 缀合物； (2) 确定结果 干扰培养的果蝇细胞中的 Smt3 缀合，例如 蛋白质定位和转录调控； (3) 确定 通过遗传研究 Smt3 结合在完整生物体中的发育作用 并逆转扰乱 Smt3 体内缀合的遗传方法， 目标是在观察到的表型和特定目标之间建立直接联系 smt3-共轭。 这些研究对人类健康有很多影响。例如，一个 最近发现的 Smt3 结合靶点是早幼粒细胞白血病 蛋白质 PML。该蛋白与 Smt3 的结合似乎可以控制其 定位到称为 PML 小体的离散核灶。最近的实验 表明果蝇细胞核可能含有类似的 Smt3 依赖性病灶。