Neuropeptide F interacts with its receptor and regulates the immune response of Sepiella japonica

In mammals, neuropeptide Y (NPY) has been recognized for its role in modulating the immune response of host. However, invertebrate neuropeptide F (NPF), as a homologous gene of NPY, has been minimally explored immunomodulatory function. In this study, NPF and NPF receptor ( NPFR ) mRNAs were significantly up-regulated in sick Sepiella japonica , and in juvenile S. japonica after bath infection of Vibrio harveyi . Subsequently, hemocytes of S. japonica were isolated, and Sj _ NPF and Sj _ NPFR mRNAs were notably induced in the hemocyte of S. japonica after Lipopolysaccharide (LPS) stimulation. Flow cytometry and pull-down assay confirmed the interaction between Sj _NPF and Sj _NPFR. Afterwards, knockdown of NPF notably decreased the mRNA expression of NPFR, Nitric oxide synthase ( NOS ) , Toll-like receptor ( TLR ) and Interleukin-17-7 ( IL-17-7 ) in the white body of S. japonica . Furthermore, after in vivo injection of NPF mature peptide, NPFR , NOS , TLR , TNF and IL-17- 7 mRNAs were significantly up-regulated in the white body of S. japonica , with TNF and IL-17- 7 mRNAs also showing significant induction in the liver of S. japonica . Interestingly, NPFR , TLR , MyD88 and IL-17- 7 mRNAs were significantly decreased in the gill of S. japonica after in vivo injection of NPF mature peptide. Finally, the colocalization of Sj _ NPF and Sj _ NOS mRNAs was observed in the dorsal area of juvenile cuttlefish using whole-mount in situ hybridization (WISH), and NO content was increased at 24 h and 48 h in the liver after in vivo injection of NPF mature peptide. Collectively, these results uncover that the Sj _NPF may interact with its receptor and mediate immune response to eliminate invasive pathogens. • Sj _ NPF and Sj _ NPFR mRNAs were significantly up-regulated after infection of Vibrio harveyi and LPS stimulation. • Flow cytometry and pull-down assay confirmed the interaction between Sj _NPF and Sj _NPFR. • Knockdown of Sj _NPF notably decreased the mRNA expression of immune-related genes in the white body of S. japonica. • These immune-related genes mRNAs were significantly up-regulated after in vivo injection of NPF mature peptide. • The colocalization of Sj _ NPF and Sj _ NOS mRNAs was observed in the dorsal area of juvenile cuttlefish using WISH.