Mechanisms of Delta-like 4 Endocytosis and Notch Activation During Blood Vessel Development

血管发育过程中 Delta-like 4 内吞作用和 Notch 激活的机制

基本信息

项目摘要

SUMMARY Endothelial cells (ECs) are the cell type responsible for the bulk of embryonic blood vessel formation, eventually leading to an estimated 100,000 miles of vasculature by adulthood. During development, new blood vessels emerge from pre-existing vasculature, a process termed angiogenesis. Notch signaling is fundamental to angiogenesis and adult blood vessel homeostasis. In the absence of Notch, blood vessels demonstrate a chronic sprouting phenotype marked by unchecked proliferation and overgrowth; this evidence and others overwhelmingly show Notch is required for blood vessel maturation and stability. When ligand-bound, the Notch intracellular domain is cleaved and translocates to the nucleus, acting as a transcription factor. Delta-like ligand 4 (DLL4) is a potent Notch ligand that binds to the extracellular domain of Notch. To elicit Notch activation the ectodomain of Notch requires a pulling force by DLL4 to expose a S2 cleavage site. Once exposed, the Notch extracellular domain is cleaved, allowing for release of the Notch intracellular domain (NICD) stimulating signaling activation. Very little is known about the post-transcriptional regulation of DLL4, and even less is understood about the mechanisms by which DLL4 exerts a sustained pulling force to activate Notch-mediated lateral inhibition. Our preliminary data describes, for the first time, how DLL4 endocytosis and anchoring to the actin cytoskeleton generates the mechanical force required to expose the S2 site of Notch. Specifically, this proposal will focus on two largely uncharacterized proteins we believe are central to force- generating DLL4 endocytosis, Eps15 homology domain binding protein 1 (EHBP1) and EH domain containing protein 2 (EHD2). Our broad hypothesis is that EHBP1 anchors EHD2 to f-actin filaments, aiding in the transendocytosis of DLL4 bound to Notch. In Aim1 we will detail how both EHBP1 and EHD2 work in combination to facilitate DLL4 endocytosis, and in doing so, mediate Notch signaling. In Aim2, we will comprehensively demonstrate that EHBP1 and EDH2 works in combination to regulate DLL4 endocytosis and downstream Notch activity during zebrafish blood vessel morphogenesis using live-imaging and CRISPR- based mutant generation. Overall, this proposal will answer several critically important questions pertaining to blood vessel development as well as provide a powerful training opportunity for undergraduate scholars.
摘要 内皮细胞(ECs)是胚胎血管形成的主要细胞类型, 最终导致成年后大约有10万英里长的血管形成。在发展过程中,新的血液 血管从预先存在的血管系统中产生,这一过程被称为血管生成。陷波信令是基本的 血管生成和成人血管动态平衡。在没有Notch的情况下,血管表现为 以无节制的增殖和过度生长为特征的慢性萌芽表型;这一证据和其他 压倒性地表明Notch是血管成熟和稳定所必需的。当与配体结合时, Notch胞内结构域被切割并移位到细胞核,作为转录因子发挥作用。类似于三角洲的 配体4(DLL4)是一种有效的Notch配体,可与Notch的胞外结构域结合。引出缺口 激活Notch的胞外结构域需要DLL4的拉力来暴露S2裂解位点。一次 暴露后,Notch胞外区被切割,允许Notch胞内区释放 (NiCd)刺激信号激活。对DLL4的转录后调控知之甚少, 对DLL4施加持续拉力以激活的机制更是知之甚少 缺口介导的侧向抑制。我们的初步数据首次描述了DLL4内吞作用和 锚定到肌动蛋白细胞骨架上会产生暴露Notch的S2部位所需的机械力。 具体地说,这项提案将专注于两种我们认为是推动- 产生DLL4内吞作用,Eps15同源结构域结合蛋白1(EHBP1)和EH结构域包含 蛋白2(EHD2)。我们的广泛假设是,EHBP1将EHD2锚定在f-肌动蛋白细丝上,帮助 结合Notch的DLL4的跨内吞作用。在Aim1中,我们将详细介绍EHBP1和EHD2如何在 结合以促进DLL4的内吞作用,并在此过程中,介导Notch信号。在AIM2中,我们将 全面证明EHBP1和EDH2联合作用调节DLL4内吞和 斑马鱼血管形态发生过程中下游Notch活性的活体成像和CRISPR- 基于突变的一代。总体而言,这项提案将回答以下几个至关重要的问题 同时也为本科生提供了强有力的培训机会。

项目成果

期刊论文数量(8)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Rab35 governs apicobasal polarity through regulation of actin dynamics during sprouting angiogenesis.
  • DOI:
    10.1038/s41467-022-32853-5
  • 发表时间:
    2022-09-08
  • 期刊:
  • 影响因子:
    16.6
  • 作者:
  • 通讯作者:
Lipidure-based micropattern fabrication for stereotyping cell geometry.
  • DOI:
    10.1038/s41598-023-47516-8
  • 发表时间:
    2023-11-22
  • 期刊:
  • 影响因子:
    4.6
  • 作者:
  • 通讯作者:
Trafficking in blood vessel development.
  • DOI:
    10.1007/s10456-022-09838-5
  • 发表时间:
    2022-08
  • 期刊:
  • 影响因子:
    9.8
  • 作者:
    Francis CR;Kushner EJ
  • 通讯作者:
    Kushner EJ
Capturing membrane trafficking events during 3D angiogenic development in vitro.
EHD2 modulates Dll4 endocytosis during blood vessel development.
  • DOI:
    10.1111/micc.12740
  • 发表时间:
    2022-01
  • 期刊:
  • 影响因子:
    2.4
  • 作者:
    Webb, Amelia M.;Francis, Caitlin R.;Judson, Rachael J.;Kincross, Hayle;Lundy, Keanna M.;Westhoff, Dawn E.;Meadows, Stryder M.;Kushner, Erich J.
  • 通讯作者:
    Kushner, Erich J.
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Erich J Kushner其他文献

Erich J Kushner的其他文献

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{{ truncateString('Erich J Kushner', 18)}}的其他基金

Polarized Protein Trafficking and Angiogenesis
极化蛋白运输和血管生成
  • 批准号:
    10363446
  • 财政年份:
    2022
  • 资助金额:
    $ 43.9万
  • 项目类别:
Polarized Protein Trafficking and Angiogenesis
极化蛋白运输和血管生成
  • 批准号:
    10539327
  • 财政年份:
    2022
  • 资助金额:
    $ 43.9万
  • 项目类别:
Mechanisms of Basement Membrane Regulation During Angiogenesis
血管生成过程中基底膜的调节机制
  • 批准号:
    10002605
  • 财政年份:
    2019
  • 资助金额:
    $ 43.9万
  • 项目类别:
Centrosomes and Cytoskeletal Mechanisms of Blood Vessel Dysfunction
血管功能障碍的中心体和细胞骨架机制
  • 批准号:
    8891096
  • 财政年份:
    2015
  • 资助金额:
    $ 43.9万
  • 项目类别:
Centrosome Over-duplication and Blood Vessel Function
中心体过度复制与血管功能
  • 批准号:
    8455123
  • 财政年份:
    2013
  • 资助金额:
    $ 43.9万
  • 项目类别:
Centrosome Over-duplication and Blood Vessel Function
中心体过度复制与血管功能
  • 批准号:
    8627974
  • 财政年份:
    2013
  • 资助金额:
    $ 43.9万
  • 项目类别:

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A novel motility system driven by two classes of bacterial actins MreB
由两类细菌肌动蛋白 MreB 驱动的新型运动系统
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Cytoplasmic Actins in Maintenance of Muscle Mitochondria
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多种植物肌动蛋白的差异表达
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Studies on how actins and microtubules are coordinated and its relevancy.
研究肌动蛋白和微管如何协调及其相关性。
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拟南芥生殖肌动蛋白的抑制
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    6655612
  • 财政年份:
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Suppression of Arabidopsis Reproductive Actins
拟南芥生殖肌动蛋白的抑制
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肌球蛋白与单体肌动蛋白的相互作用
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STRUCTURE/INTERACTIONS OF ACTINS AND ACTIN-BINDING PROTEIN
肌动蛋白和肌动蛋白结合蛋白的结构/相互作用
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    6316669
  • 财政年份:
    2000
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