O-Glycosylation of Epidermal Growth Factor-like Modules
表皮生长因子样模块的 O-糖基化
基本信息
- 批准号:8197516
- 负责人:
- 金额:$ 62.64万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2001
- 资助国家:美国
- 起止时间:2001-01-01 至 2013-09-15
- 项目状态:已结题
- 来源:
- 关键词:AblationAcute T Cell LeukemiaAffectAlagille SyndromeArterial DisorderBiological AssayCADASILCarbohydratesCell NucleusCell Surface ReceptorsCell membraneCellsCerebrumCryoelectron MicroscopyDataDefectDevelopmentDiseaseDrosophila genusDysostosesEnzymesEpidermal Growth FactorEventExtracellular DomainFamilyFucoseGenesGeneticGenetic TranscriptionGlucoseGlucosyltransferaseHealthHomologous GeneHumanHuman PathologyInfarctionLeadLeukoencephalopathyLigand BindingLigandsLinkMalignant NeoplasmsMalignant neoplasm of cervix uteriMammalian CellMapsMediatingMethodologyMethodsModificationMolecularMolecular ConformationMusMutationN-AcetylglucosaminyltransferasesNatureNotch Signaling PathwayPeptide HydrolasesPhenotypePlayPolysaccharidesPost-Translational Protein ProcessingProteinsProteolysisRoleShapesSignal TransductionSiteSmall Interfering RNAStagingStructureSystemTemperatureTestingTrisaccharidesVascular DiseasesXyloseanalytical ultracentrifugationbasecancer typedesigndevelopmental diseaseflexibilityflyglycosylationhuman diseaseinnovationmelanomanotch proteinprotein structurepublic health relevanceresearch studysecretasesugarxylosyltransferase
项目摘要
DESCRIPTION (provided by applicant): Notch receptors play an essential role in numerous stages of development, and deregulation of Notch signaling leads to a variety of human pathologies, including cancers, vascular disorders, and developmental disorders. The Notch extracellular domain (ECD) contains up to 36 tandem epidermal growth factor-like (EGF) repeats, many of which are modified by two unusual forms of glycosylation: O-fucose and O-glucose. O-Fucose modifications are essential for Notch function. Genetic ablation of the enzyme that adds O-fucose to EGF repeats causes severe Notch-like phenotypes in either mice or flies, and elongation of O-fucose by the Fringe family of ss3-N-acetylglucosaminyltransferases modulates Notch signaling. Fringe functions by altering the interaction between Notch and its ligands: Delta or Serrate/Jagged. Interestingly, Fringe modification increases Notch-Delta interactions while inhibiting Notch-Serrate/Jagged interactions. We have recently shown that O-glucose modifications are also essential for Notch function. Mutations in the gene encoding the enzyme responsible for addition of O-glucose to EGF repeats (protein O-glucosyltransferase, or Rumi), cause a temperature sensitive Notch-like phenotype in flies. Although both O-fucose and O-glucose modifications are required for Notch function, the molecular details for how they mediate their effects are not known. Our hypothesis is that the O-fucose and O-glucose glycans affect Notch function by affecting the conformation of the Notch ECD. This hypothesis is based on exciting recent structural studies suggesting that the Notch ECD has regions of flexibility that are evolutionarily conserved. In the first aim we will analyze how Fringe modification alters Notch-ligand binding. Using quantitative mass spectral methods, we will map Fringe modification sites, and correlate these modifications with changes in Notch-ligand binding. These studies will tell us which regions of the Notch ECD are involved in regulating Notch-ligand binding. We will examine the possibility that some regions of the Notch ECD inhibit ligand binding, and we will test whether Fringe modulates this inhibitory activity. Finally, we will examine whether Fringe induces changes in the shape of the Notch ECD using protease sensitivity, analytical ultracentrifugation, and an innovative cryoEM method. In the second aim, we will identify and characterize mammalian homologue of Rumi, as well as the xylosyltransferases that elongate O-glucose with xyloses. The final aim is designed to test whether elimination of Rumi (or the xylosyltransferases identified in Aim 2) in mammalian cells (using siRNA strategies) causes a loss of Notch activity as was seen in flies. In addition, we will examine whether loss of O-glucose causes conformational changes in the Notch ECD using the same methodologies described in Aim 1 for the effects of Fringe on Notch. These experiments will provide molecular details for how these unusual carbohydrate modifications alter Notch function. Public Health Relevance: Defects in the Notch signaling pathway lead to a variety of human pathologies, including several types of cancer, vascular disorders, and developmental disorders. Notch is regulated at numerous levels, including by glycosylation (modification of proteins by sugars). Our studies are aimed at understanding how glycosylation affects Notch activity so that we can take advantage of its ability to regulate Notch to design potential therapies for Notch-related diseases.
描述(由申请人提供):Notch受体在许多发育阶段中发挥重要作用,Notch信号传导的失调导致多种人类病理学,包括癌症、血管疾病和发育障碍。Notch细胞外结构域(ECD)包含多达36个串联的表皮生长因子样(EGF)重复序列,其中许多被两种不寻常的糖基化形式修饰:O-岩藻糖和O-葡萄糖。O-岩藻糖修饰对于Notch功能是必不可少的。将O-岩藻糖添加到EGF重复序列的酶的遗传消融在小鼠或苍蝇中引起严重的Notch样表型,并且β 3-N-乙酰葡糖胺转移酶的Fringe家族对O-岩藻糖的延长调节Notch信号传导。Fringe通过改变Notch及其配体之间的相互作用发挥作用:Delta或Serrate/Jagged。有趣的是,条纹修饰增加Notch-Delta相互作用,同时抑制Notch-Serrate/Jagged相互作用。我们最近表明,O-葡萄糖修饰对Notch功能也是必不可少的。编码负责将O-葡萄糖添加到EGF重复序列(蛋白质O-葡萄糖基转移酶或Rumi)的酶的基因突变会导致果蝇中的温度敏感性Notch样表型。虽然O-岩藻糖和O-葡萄糖修饰都是Notch功能所必需的,但它们如何介导其作用的分子细节尚不清楚。我们的假设是O-岩藻糖和O-葡萄糖聚糖通过影响Notch ECD的构象来影响Notch功能。这一假设是基于令人兴奋的最近的结构研究表明,Notch ECD具有进化保守的灵活性区域。在第一个目标中,我们将分析条纹修饰如何改变Notch-配体结合。使用定量质谱方法,我们将绘制条纹修饰位点,并将这些修饰与Notch-配体结合的变化相关联。这些研究将告诉我们Notch ECD的哪些区域参与调节Notch-配体结合。我们将研究Notch ECD的某些区域抑制配体结合的可能性,并且我们将测试Fringe是否调节这种抑制活性。最后,我们将研究是否条纹诱导的Notch ECD的形状变化,使用蛋白酶的敏感性,分析ultracenthesis,和一个创新的cryoEM方法。在第二个目标中,我们将确定和表征哺乳动物同源的鲁米,以及木糖基转移酶,延长O-葡萄糖与木糖。最终目的是测试哺乳动物细胞中Rumi(或Aim 2中鉴定的木糖基转移酶)的消除(使用siRNA策略)是否会导致如在果蝇中所见的Notch活性丧失。此外,我们将使用目标1中描述的用于Fringe对Notch的影响的相同方法来检查O-葡萄糖的损失是否引起Notch ECD中的构象变化。这些实验将为这些不寻常的碳水化合物修饰如何改变Notch功能提供分子细节。公共卫生相关性:Notch信号通路的缺陷导致多种人类病理学,包括几种类型的癌症、血管疾病和发育障碍。Notch在许多水平上受到调节,包括通过糖基化(糖对蛋白质的修饰)。我们的研究旨在了解糖基化如何影响Notch活性,以便我们可以利用其调节Notch的能力来设计Notch相关疾病的潜在疗法。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Robert S. Haltiwanger其他文献
<em>O</em>-Fucose modification is essential for patterning mesoderm in the mouse embryo
- DOI:
10.1016/j.ydbio.2008.05.422 - 发表时间:
2008-07-15 - 期刊:
- 影响因子:
- 作者:
Jianguang Du;Hideyuki Takeuchi;Christina Leonhard;Malgosia Dlugosz;Robert S. Haltiwanger;Bernadette C. Holdener - 通讯作者:
Bernadette C. Holdener
13-P011 Restriction of EMT within the primitive streak and correct patterning of the mesoderm requires Pofut2
- DOI:
10.1016/j.mod.2009.06.484 - 发表时间:
2009-08-01 - 期刊:
- 影响因子:
- 作者:
Jianguang Du;Christina L. Leonhard-Melief;Hideyuki Takeuchi;Kenneth R. Shroyer;Malgosia Dlugosz;Robert S. Haltiwanger;Bernadette C. Holdener - 通讯作者:
Bernadette C. Holdener
FUT10 and FUT11 are protein O-fucosyltransferases that modify protein EMI domains
FUT10 和 FUT11 是修饰蛋白质 EMI 结构域的蛋白质 O-岩藻糖基转移酶
- DOI:
10.1038/s41589-024-01815-x - 发表时间:
2025-01-07 - 期刊:
- 影响因子:13.700
- 作者:
Huilin Hao;Youxi Yuan;Atsuko Ito;Benjamin M. Eberand;Harry Tjondro;Michelle Cielesh;Nicholas Norris;Cesar L. Moreno;Joshua W. C. Maxwell;G. Gregory Neely;Richard J. Payne;Melkam A. Kebede;Ramona J. Bieber Urbauer;Freda H. Passam;Mark Larance;Robert S. Haltiwanger - 通讯作者:
Robert S. Haltiwanger
Analysis of the Healthy Platelet Proteome Identifies a New Form of Domain-Specific emO-/emFucosylation
健康血小板蛋白质组的分析确定了一种新形式的域特异性表情符/纤维糖基化
- DOI:
10.1016/j.mcpro.2024.100717 - 发表时间:
2024-02-01 - 期刊:
- 影响因子:5.500
- 作者:
Callum B. Houlahan;Yvonne Kong;Bede Johnston;Michelle Cielesh;The Huong Chau;Jemma Fenwick;Paul R. Coleman;Huilin Hao;Robert S. Haltiwanger;Morten Thaysen-Andersen;Freda H. Passam;Mark Larance - 通讯作者:
Mark Larance
Novel antibodies detect nucleocytoplasmic O-fucose in protist pathogens, cellular slime molds, and plants
新型抗体检测原生生物病原体、细胞黏菌和植物中的核质 O-岩藻糖
- DOI:
10.1128/msphere.00945-24 - 发表时间:
2025-02-03 - 期刊:
- 影响因子:3.100
- 作者:
Megna Tiwari;Elisabet Gas-Pascual;Manish Goyal;Marla Popov;Kenjiroo Matsumoto;Marianne Grafe;Ralph Gräf;Robert S. Haltiwanger;Neil Olszewski;Ron Orlando;John C. Samuelson;Christopher M. West - 通讯作者:
Christopher M. West
Robert S. Haltiwanger的其他文献
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{{ truncateString('Robert S. Haltiwanger', 18)}}的其他基金
Glycosylation of Thrombospondin Type 1 Repeats
血小板反应蛋白 1 型重复序列的糖基化
- 批准号:
7266505 - 财政年份:2007
- 资助金额:
$ 62.64万 - 项目类别:
Glycosylation of Thrombospondin Type 1 Repeats
血小板反应蛋白 1 型重复序列的糖基化
- 批准号:
7556767 - 财政年份:2007
- 资助金额:
$ 62.64万 - 项目类别:
Glycosylation of Thrombospondin Type 1 Repeats
血小板反应蛋白 1 型重复序列的糖基化
- 批准号:
8018543 - 财政年份:2007
- 资助金额:
$ 62.64万 - 项目类别:
Glycosylation of Thrombospondin Type 1 Repeats
血小板反应蛋白 1 型重复序列的糖基化
- 批准号:
7759150 - 财政年份:2007
- 资助金额:
$ 62.64万 - 项目类别:
Glycosylation of Thrombospondin Type 1 Repeats
血小板反应蛋白 1 型重复序列的糖基化
- 批准号:
7357473 - 财政年份:2007
- 资助金额:
$ 62.64万 - 项目类别:
Gordon Research Conference on Glycobiology 2005/2007
戈登糖生物学研究会议 2005/2007
- 批准号:
6945432 - 财政年份:2004
- 资助金额:
$ 62.64万 - 项目类别:
Gordon Research Conference on Glycobiology 2005/2007
戈登糖生物学研究会议 2005/2007
- 批准号:
7023729 - 财政年份:2004
- 资助金额:
$ 62.64万 - 项目类别:
Gordon Research Conference on Glycobiology 2005/2007
戈登糖生物学研究会议 2005/2007
- 批准号:
6887517 - 财政年份:2004
- 资助金额:
$ 62.64万 - 项目类别:
O-Glycosylation of Epidermal Growth Factor-like Motifs
表皮生长因子样基序的 O-糖基化
- 批准号:
9102203 - 财政年份:2001
- 资助金额:
$ 62.64万 - 项目类别:
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