The yellow catfish (Pelteobagrus fulvidraco) is an economic fish with a large breeding scale, and diseases have led to huge economic losses. Tumor necrosis factor receptor-associated factors (TRAFs) are a class of intracellular signal transduction proteins that play an important role in innate and adaptive immune responses by mediating NF-κB, JNK and MAPK signaling pathways. However, there are few studies on the TRAF gene family in yellow catfish. In this study, the open reading frame (ORF) sequences of TRAF1, TRAF2a, TRAF2b, TRAF3, TRAF4a, TRAF4b, TRAF5, TRAF6 and TRAF7 genes were cloned and identified in yellow catfish. The ORF sequences of the nine TRAF genes of yellow catfish (Pf_TRAF1-7) were 1413–2025 bp in length and encoded 470–674 amino acids. The predicted protein structures of Pf_TRAFs have typically conserved domains compared to mammals. The phylogenetic relationships showed that TRAF genes are conserved during evolution. Gene structure, motifs and syntenic analyses of TRAF genes showed that the exon–intron structure and conserved motifs of TRAF genes are diverse among seven vertebrate species, and the TRAF gene family is relatively conserved evolutionarily. Among them, TRAF1 is more closely related to TRAF2a and TRAF2b, and they may have evolved from a common ancestor. TRAF7 is quite different and distantly related to other TRAFs. Real-time quantitative PCR (qRT-PCR) results showed that all nine Pf_TRAF genes were constitutively expressed in 12 tissues of healthy yellow catfish, with higher mRNA expression levels in the gonad, spleen, brain and gill. After infection with Edwardsiella ictaluri, the expression levels of nine Pf_TRAF mRNAs were significantly changed in the head kidney, spleen, gill and brain tissues of yellow catfish, of which four genes were down-regulated and one gene was up-regulated in the head kidney; four genes were up-regulated and four genes were down-regulated in the spleen; two genes were down-regulated, one gene was up-regulated, and one gene was up-regulated and then down-regulated in the gill; one gene was up-regulated, one gene was down-regulated, and four genes were down-regulated and then up-regulated in the brain. These results indicate that Pf_TRAF genes might be involved in the immune response against bacterial infection. Subcellular localization results showed that all nine Pf_TRAFs were found localized in the cytoplasm, and Pf_TRAF2a, Pf_TRAF3 and Pf_TRAF4a could also be localized in the nucleus, uncovering that the subcellular localization of TRAF protein may be closely related to its structure and function in cellular mechanism. The results of this study suggest that the Pf_TRAF gene family plays important roles in the immune response against pathogen invasion and will provide basic information to further understand the roles of TRAF gene against bacterial infection in yellow catfish.

Neuropeptide F (NPF), an invertebrate homolog of vertebrate neuropeptide Y (NPY), exerts pleiotropic functions through its interaction with the G protein-coupled receptor (GPCR) neuropeptide F receptor (NPFR). However, the role of the NPF/NPFR system in the Chinese common cuttlefish Sepiella japonica Sasaki, 1929—a commercially and scientifically important cephalopod species in East China Sea aquaculture—remains unclear. In the present study, SjNPF/SjNPFR genes were cloned from S. japonica. Multiple alignments demonstrated that SjNPF/SjNPFR exhibited a high identity with that of other cephalopods. Spatio-temporal expression patterns revealed that SjNPF and SjNPFR transcripts were relatively highly expressed in the central nervous and digestive systems across all developmental stages. In situ hybridization (ISH) monitored clear and stable positive signals of SjNPF and SjNPFR mRNA at the junction of the subvertical lobe and the vertical lobe, as well as in the brachial lobe, pedal lobe and the palleovisceral lobe. Subcellular localization studies showed that SjNPF was primarily localized in the cytoplasm, whereas SjNPFR was membrane-localized. Moreover, under feeding-regulatory conditions (5-day starvation followed by 3-day refeeding), mRNA expression levels of SjNPF and SjNPFR in the treated group were positively correlated with starvation and negatively correlated with refeeding. These findings provide valuable insights for future investigations into the pleiotropic functional roles of the NPF/NPFR system in S. japonica and the peptidergic regulation of this system in cephalopods.

5-Hydroxytryptamine (5-HT) plays a vital role in the reproductive process of vertebrates and is also present in many invertebrates. The cDNA of the Sepiella japonica 5-HT₆ receptor (Sj5-HT₆r) was first cloned by RACE (Rapid Amplification of cDNA Ends). The length was 1450 bp, and the predicted open reading frame (ORF) was 1116 bp, which encoded 371 amino acids. Sequence characteristics analysis showed that Sj5-HT₆r shares a high degree of identity with 5-HT₆r from other cephalopods and forms a sister branch to bivalves. Subcellular localization showed that Sj5-HT₆r protein was localized on the HEK293T cell membrane surface. Quantitative Real-time PCR (qPCR) analysis demonstrated that Sj5-HT₆r was highly expressed in reproductive organs of both sexes. In particular, transcripts with significant expression were observed at stage III of female gonadal development in tissues of the ovary and nidamental gland, and at stage IV in tissues of the accessory nidamental gland. In situ hybridization (ISH) experiment results indicated that Sj5-HT₆r mRNA was primarily distributed in all regions of the optic lobes except the plexiform zone. These results may provide a basis for the future exploration of the reproductive regulation of 5-HT and 5-HT₆ receptors in S. japonica.