Smt3-conjugation in cell biology and development

细胞生物学和发育中的 Smt3 缀合

• 批准号: 6657425
• 负责人: ALBERT J COUREY
• 金额: $ 18.62万
• 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 2005-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6657425
• 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）：类似泛素的蛋白质正在接受 由于他们最近发现的角色在多种多样而引起的关注不断增加 生物过程。一种这样的蛋白质，称为SMT3（也称为Sumo或 Sentrin），具有广泛的可疑功能，包括功能 转录调节，核转运和细胞因子。像泛素一样， SMT3通过异肽链接与目标蛋白共价连接到目标蛋白 赖氨酸残留物。这些目标包括转录因子， 核孔复合物和隔丁复合物的成分。而很多 已经确定了SMT3结合靶标，对SMT3功能的研究 后生动物在很大程度上仅限于细胞培养分析。这个建议 描述了对生化，细胞生物学和遗传分析的组合 果蝇中的SMT3结合，目的是确定功能 这一过程在发展中的生物体中的作用。 具体目的是：（1）确定在 果蝇通过在体内表达标记的SMT3形式，然后使用标签 从蛋白质提取物中纯化smt3结合； （2）确定结果 在培养的果蝇细胞中扰动SMT3结合在诸如等过程中 蛋白质定位和转录调节； （3）确定 通过使用遗传 和反向遗传方法在体内扰动SMT3结合的方法 在观察到的表型和特定目标之间建立直接联系的目标 SMT3结合。 这些研究对人类健康有许多影响。例如，一个 最近发现的SMT3结合的靶标的是promyelocytic白血病 蛋白质PML。该蛋白与SMT3的结合似乎控制了其 定位以离散的核灶称为PML体。最近的实验 表明果蝇核可能包含相似的SMT3依赖性灶。