Proteomics For Liver Of Pseudosciaena Crocea Induced By Poly(I:C) And Identification Of Two Immune Genes

Large yellow croaker (Pseudosciana crocea) is a proper economic fishes in China. It is welcomed because of its good taste and medical values. Artificial aquaculture of large yellow croaker was developed enormously after breeding and acquaculture technology was handled. Viral diseases, which bring huge damage for acquaculture, were induced by large scale acquaculture and worsing water condition. In the current study, we analyzed the liver samples of the control and experimental groups induced by Poly(I:C) of Pseudosciaena crocea for the understanding of anti-disease mechanism in fish on the level of molecule by using high scale and high resolution of proteomic methodologies. 30 differential spots in liver sample were characterized.fd These changed spots were analyzed by PMF (peptide mass fingerprint) through MALDI-TOF-MS firstly. Furthermore, 16 differential spots were analyzed by MALDI-TOF-TOF and 13 were identified, which we got the identified peptides. In study on antidisease mechanism of Pseudosciaena crocea, we inquired about the functions of these diffrerential spots, indicating that 13.3%, 40%, 10%, 13.3%,13.3% and 10% spots are about immunine proteins, metabolized enzymes, regulation proteins, transcribed/ translation proteins, structural proteins and no functionial proteins, respectively.In human cells, interferon-inducible transmembrane protein 1(IFITM1) is a component of protein complexes involved in homotypic adhesion and the transduction of antiproliferative signals. Here, we reported the cloning of an IFITM1 homologue from the spleen of large yellow croaker Pseudosciaena crocea (LycIFITM1). The complete cDNA of LycIFITM1 is 734 nucleotides (nt) encoding a protein of 124amino acids (aa), with a putative molecular weight of 13.6 kDa. The deduced LycIFITM1 protein is significantly homologous to interferon-inducible transmembrane proteins (IFITMs) in mammals and fish, and has the typical structural features of IFITMs, including two transmembrane domains (residues 43-63 and 90-112, respectively) and one intracellular domain between them (residues 64-89), as well as one conserved protein kinase C (PKC) phosphorylation site (residues 65-67, SIK). Phylogenetic analysis showed that LycIFITM1 formed a cluster with fish IFITM, reflecting a relative distant evolutionary relationship from mammals. LycIFITM1 gene was constitutively expressed in various tissues examined, such as gills, intestine, liver, kidney, heart, spleen, muscle and blood. Upon induction with poly(I:C), LycIFITM1 gene expression was obviously up-regulated in gills, kidney, heart and spleen at 24h after stimulation, suggesting that LycIFITM1 may be involved in the immune response induced by poly(I:C). Time course analysis using real-time PCR showed that the mRNA levels of LycIFITM1 in spleen and kidney were quickly up-regulated by poly(I:C) and reached the peak at 24h post-induction (48.7-and 280.4-fold mRNA increases in spleen and kidney, respectively). The results suggest that the IFITM1 homologue from large yellow croaker may represent a member of IFITMs family in fish.In the present study, we reported the cloning of a CXCL12 chemokine gene homologue from large yellow croaker Pseudosciaena crocea (LycCXCL12). The complete cDNA of LycCXCL12 is 678 nucleotides (nt) encoding a protein of 97 amino acids (aa), with a putative molecular weight of 11.1 kDa. The deduced LycCXCL12 contains a 22-aa signal peptide and a 75- aa mature polypeptide,which possesses the typical arrangement of four cysteines as found in other known CXC chemokines (C31, C33, C56and C71). Phylogenetic analysis showed that LycCXCL12 was more closely related to the CXCL12 subgroup than to any other CXC chemokine subgroups. LycCXCL12 gene was constitutively expressed in all tissues examined, such as gills, intestine, liver, kidney, heart, spleen, muscle and blood. Upon induction with poly(I:C) or inactivated trivalent bacterial vaccine, LycCXCL12 gene expression was significantly up-regulated in gills, liver, kidney, spleen and blood at 24h after stimulation, and bacterial vaccine was more potent than poly(I:C) in up-regulating LycCXCL12 expression in these tissues. Time course analysis using real-time PCR showed that LycCXCL12 gene expression in spleen, kidney and gills was quickly up-regulated by either poly(I:C) or bacterial vaccine, and reached the peak level at 12 h (in spleen and kidney) or 24h (in gills) post-induction by poly(I:C) and at 24h (in kidney ) or 48h (in gills and spleen) post-induction by bacterial vaccine, respectively. These results suggest that LycCXCL12 may have a role in inflammatory response as well as homeostatic process.