Litopenaeus vannamei Notch interacts with COP9 signalosome complex subunit 1 (CNS1) to negatively regulate the NF-κB pathway

Abstract 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.