Journal of Proteomics ( IF 2.8 ) Pub Date : 2020-12-10 , DOI: 10.1016/j.jprot.2020.104074 Weiling Zhao 1 , Zhihong Zheng 1 , Jude Juventus Aweya 1 , Fan Wang 1 , Shengkang Li 1 , Tran Ngoc Tuan 1 , Defu Yao 1 , Yueling Zhang 2
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Notch signaling pathway is a highly evolutionary conserved signaling pathway, which modulates many biological processes such as cell differentiation, tissue development and immune response. Our previous study revealed that Litopenaeus vannamei Notch (LvNotch) was involved in immune response by regulating reactive oxygen species (ROS) production in hemocytes. However, the immune regulatory networks mediated by LvNotch remain unclear in shrimp. In this study, 21 proteins that potentially interact with LvNotch were identified by GST pull-down and liquid chromatography tandem mass spectrometry (LC-MS/MS) analyses. Among these proteins, COP9 signalosome complex subunit 1 (CSN1) was chosen for further studies due to its putative role in immune response. The interaction between LvNotch and LvCSN1 was confirmed by Far-Western blot and GST pull-down analyses. In vivo knockdown of LvNotch resulted in an increase in LvCSN1 expression in hemocytes, which suggest that the COP9 signalosome complex might be negatively regulated by LvNotch. In addition, in vivo silencing of LvNotch upregulated the expression of LvDorsal, LvTNFSF and LvCrustin2 (NF-κB pathway related-genes), while their expression decreased after LvCSN1 depletion. Collectively, the current results indicate that LvNotch negatively regulates the NF-κB pathway by modulating LvCSN1 in shrimp.
Significance
Although the Notch signaling pathway has been implicated in the regulation of immune response in vertebrates and invertebrates, the functions and immune-related interacting networks of Notch in shrimp immune response remain unknown. In this study, twenty-one proteins including COP9 signalosome complex subunit 1 (CSN1) were identified as potential interacting partners of LvNotch. Further analysis revealed that LvNotch negatively regulates the NF-κB pathway by binding to CSN1 and modulating its expression. These findings for the first time suggest that the Notch signaling pathway has cross-talk with the NF-κB pathway in shrimp as part of the immune response.
中文翻译：
凡纳滨对虾Notch与COP9信号体复合物亚基1（CNS1）相互作用以负调节NF-κB通路
Notch信号传导途径是高度进化的保守信号传导途径，可调节许多生物学过程，例如细胞分化，组织发育和免疫反应。我们先前的研究表明，凡纳滨对虾（Lv Notch）通过调节血细胞中的活性氧（ROS）产生而参与免疫应答。但是，虾中由Lv Notch介导的免疫调节网络仍不清楚。在这项研究中，有21种可能与Lv相互作用的蛋白通过GST下拉和液相色谱串联质谱（LC-MS / MS）分析鉴定缺口。在这些蛋白质中，由于COP9信号体复合物亚基1（CSN1）在免疫应答中的假定作用，因此被选择用于进一步研究。Lv Notch和Lv CSN1之间的相互作用已通过Far-Western印迹和GST下拉分析得到了证实。体内敲除Lv Notch导致血细胞中Lv CSN1表达增加，这表明COP9信号复合体可能受Lv Notch负调节。此外，Lv Notch的体内沉默上调了Lv的表达背，Lv TNFSF和Lv Crustin2（NF-κB通路相关基因），而在Lv CSN1耗尽后它们的表达下降。总体而言，当前结果表明，Lv Notch通过调节虾中的Lv CSN1负调节NF-κB途径。
意义
尽管Notch信号通路与脊椎动物和无脊椎动物免疫应答的调节有关，但是Notch在虾免疫应答中的功能和与免疫相关的相互作用网络仍然未知。在这项研究中，包括COP9信号体复合物亚基1（CSN1）在内的二十一种蛋白被鉴定为Lv Notch的潜在相互作用伴侣。进一步的分析表明，Lv Notch通过与CSN1结合并调节其表达来负调节NF-κB通路。这些发现首次表明，Notch信号通路与虾中的NF-κB通路发生串扰，这是免疫反应的一部分。
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