Integrated transcriptome analysis of immunological responses in the pearl sac of the triangle sail mussel (Hyriopsis cumingii) after mantle implantation.

For pearl culture of bivalve Hyriopsis cumingii, implantation of the sabio may cause nucleus discharge and increased host death rates. We performed a transcriptome analysis of the pearl sac of H. cumingii for 30 days after mantle implantation; 293863 unigenes were obtained, and 27176 unigenes were identified using nr, nt, KO, Swiss-Prot, Pfam, GO, and KOG databases. We detected 4878 differentially expressed genes (DEGs) through pairwise comparisons. We speculated that the physical condition of the recipient mussels returned to normal in about one month; the period was divided into six vital phases (0, 2 h-6 h, 12 h-24 h, 48 h to 7 days, 14 days and 30 days) on the basis of the overall similarities in DEGs. We compared the DEGs between time points and identified key immune-related genes. Our findings provide information on the immunological reactions induced by implantation in pearl mussels.

Identification of tumor necrosis factor receptor-associated factor 6 in the pearl mussel Hyriopsis cumingii and its involvement in innate immunity and pearl sac formation.

Tumor necrosis factor receptor-associated factor 6 (TRAF6) acts as a central intracellular signal adapter molecule that mediates the tumor necrosis factor receptor superfamily and the interleukin-1 receptor/Toll-like receptor family in vertebrates and invertebrates. In the present study, HcTRAF6, a molluscan homologue of TRAF6 from Hyriopsis cumingii, has been cloned and identified. The entire open reading frame of HcTRAF6 was found to comprise a 1965-bp region that encodes a predicted protein of 654 amino acids, which contains conserved characteristic domains including a RING domain, two TRAF-type zinc finger domains, a typical coiled coil and the MATH domain. Phylogenetic analysis revealed that HcTRAF6 was aggregated closely with CsTRAF6 from Cyclina sinensis in the invertebrate cluster of mollusks. Further, qRT-PCR analysis showed that HcTRAF6 mRNA was extensively distributed in mussel tissues with a high expression in gills. After immune stimulation with Aeromonas hydrophila and lipopolysaccharides, the transcription of HcTRAF6 was obviously induced in the gills and hemocytes. In addition, significant fluctuation in HcTRAF6 expression was observed in the pearl sac, gills and hemocytes after mantle implantation. These findings confirmed its role in the alloimmune response. Dual-luciferase reporter assay showed that over-expression of HcTRAF6 could enhance the activity of the NF-κB reporter in a dose-dependent manner. Further, the RNA interference showed that the up-regulation of antimicrobial peptides in anti-bacterial infection was strongly suppressed in HcTRAF6-silenced mussels and that depletion of HcTRAF inhibited the elimination of A. hydrophila. All these findings together prove that HcTRAF6 functions as an efficient regulator in innate immune mechanisms against invading pathogens and the alloimmune mechanism after mantle implantation in H. cumingii.