Endothelial Nitric Oxide Synthase Regulates Lymphatic Valve Specification By Controlling β - catenin Signaling During Embryogenesis.
Endothelial Nitric Oxide Synthase Regulates Lymphatic Valve Specification By Controlling β - catenin Signaling During Embryogenesis.
复制标题
内皮一氧化氮合酶通过控制胚胎发生过程中的β-连环蛋白信号传导来调节淋巴瓣膜规格。
DOI：
10.1101/2023.04.10.536303
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发表时间：
2023
期刊：
影响因子：
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通讯作者：
Scallan,JoshuaP
中科院分区：
文献类型：
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作者：
Iyer,Drishya;Mastrogiacomo,Diandra;Li,Kunyu;Banerjee,Richa;Yang,Ying;Scallan,JoshuaP
背景：淋巴阀在保证淋巴的单向运输中起着关键作用。淋巴阀丧失或功能失调与多种疾病有关，包括淋巴水肿、淋巴畸形、肥胖和回肠炎。在胚胎发生过程中，淋巴阀首先根据振荡淋巴流触发的机械转导信号通路而发育。在血管中，eNOS（基因名称：Nos3）是一种已知的剪切应力信号效应因子，但其在淋巴阀发育中的作用尚不清楚。方法我们使用全球Nos3−/−小鼠和培养的hdLECs来研究eNOS在需要振荡剪切应力（OSS）信号的淋巴阀发育中的作用。结果我们的数据显示，淋巴阀特异性细胞簇减少了45%，eNOS蛋白的缺失抑制了β-catenin的激活及其核易位。基因敲除或敲低eNOS导致体内和体外β-catenin靶蛋白下调。然而，药物抑制NO的产生并没有再现这些效果。共免疫沉淀和邻近连接实验显示eNOS直接与β-catenin结合，OSS增强了它们的结合。最后，Foxo1基因的基因消融增强了FOXC2的表达，部分挽救了eNOS基因敲除中瓣膜规范的缺失。总之，我们证明了eNOS在调节淋巴阀规范中的一种新的、不依赖于一氧化氮的作用，并提出了eNOS直接结合β-catenin调节其核易位从而调节其转录活性的机制。
BackgroundLymphatic valves play a critical role in ensuring unidirectional lymph transport. Loss of lymphatic valves or dysfunctional valves are associated with several diseases including lymphedema, lymphatic malformations, obesity, and ileitis. Lymphatic valves first develop during embryogenesis in response to mechanotransduction signaling pathways triggered by oscillatory lymph flow. In blood vessels, eNOS (gene name: Nos3) is a well characterized shear stress signaling effector, but its role in lymphatic valve development remains unexplored.MethodsWe used global Nos3−/− mice and cultured hdLECs to investigate the role of eNOS in lymphatic valve development, which requires oscillatory shear stress (OSS) signaling.ResultsOur data reveal a 45% reduction in lymphatic valve specification cell clusters and that loss of eNOS protein inhibited activation of β-catenin and its nuclear translocation. Genetic knockout or knockdown of eNOS led to downregulation of β-catenin target proteins in vivo and in vitro. However, pharmacological inhibition of NO production did not reproduce these effects. Coimmunoprecipitation and proximity ligation assays reveal that eNOS directly binds to β-catenin and their binding is enhanced by OSS. Finally, genetic ablation of the Foxo1 gene enhanced FOXC2 expression and partially rescued the loss of valve specification in the eNOS knockouts.ConclusionIn conclusion, we demonstrate a novel, nitric oxide-independent role for eNOS in regulating lymphatic valve specification and propose a mechanism by which eNOS directly binds β-catenin to regulate its nuclear translocation and thereby transcriptional activity.