TNF-Receptor-Associated Factor 3 in Litopenaeus vannamei Restricts White Spot Syndrome Virus Infection Through the IRF-Vago Antiviral Pathway

Abstract

Tumor necrosis factor receptor (TNFR)-associated factors (TRAFs) are vital signaling adaptor proteins for the innate immune response and are involved in many important pathways, such as the NF-κB-and interferon regulatory factor (IRF)-activated signaling pathways. In this study, the TRAF3 ortholog from the shrimp Litopenaeus vannamei (LvTRAF3) was cloned and characterized. LvTRAF3 has a transcript of 3,865 bp, with an open reading frame (ORF) of 1,002 bp and encodes a polypeptide of 333 amino acids, including a conserved TRAF-C domain. The expression of LvTRAF3 in the intestine and hemocyte was up-regulated in response to poly (I:C) challenge and white spot syndrome virus (WSSV) infection. RNAi knockdown of LvTRAF3 in vivo significantly increased WSSV gene transcription, viral loads, and mortality in WSSV-infected shrimp. Next, we found that LvTRAF3 was not able to induce the activation of the NF-κB pathway, which was crucial for synthesis of antimicrobial peptides (AMPs), which mediate antiviral immunity. Specifically, in dual-luciferase reporter assays, LvTRAF3 could not activate several types of promoters with NF-κB binding sites, including those from WSSV genes (wsv069, wsv056, and wsv403), Drosophila AMPs or shrimp AMPs. Accordingly, the mRNA levels of shrimp AMPs did not significantly change when TRAF3 was knocked down during WSSV infection. Instead, we found that LvTRAF3 signaled through the IRF-Vago antiviral cascade. LvTRAF3 functioned upstream of LvIRF to regulate the expression of LvVago4 and LvVago5 during WSSV infection in vivo. Taken together, these data provide experimental evidence of the participation of LvTRAF3 in the host defense to WSSV through the activation of the IRF-Vago pathway but not the NF-κB pathway.

Prx4 acts as DAMP in shrimp, enhancing bacterial resistance via the toll pathway and prophenoloxidase activation

“…Previous studies have demonstrated that an interferon (IFN)-like protein, Lv Vago5 is transcriptionally regulated by Lv IRF and it plays a key role in shrimp response following VP AHPND infection. 24 , 25 Consistent with Lv IRF expression, a remarkable upregulation of LvVago5 at the same time point following r Lv Prx4 injection was observed. These results indicate that Lv Prx4 could trigger the Toll pathway-mediated AMP production as well as the IRF-Vago pathway.…”

“…Lv Dorsal controls the transcription of antimicrobial peptide penaeidin 4 ( Lv PEN4) 48 in shrimp, that was found to be upregulated following r Lv Prx4 injection. With respect to the IRF/Vago pathway where the Lv IRF can promote the expression of the IFN-stimulated response element (ISRE)-containing promoter to trigger IFNs such as Lv Vagos, 23 , 24 increased expression of Lv IRF and Lv Vago5 following r Lv Prx4 injection was demonstrated, indicating an immune modulating role for Lv Prx4. Lv Vago5 itself has been demonstrated to participate in antibacterial response through regulation of the Toll/IMD pathway/ Lv PEN4, JAK/STAT pathway and prophenoloxidase-activating system.…”

Cloning and characterization of the LvCTL genes encoding C-type lectin from white-leg shrimp (Litopenaeus vannamei)

“…Tissue expression analysis showed LvCTL4 was distributed with high levels in the gills, intestine, epithelium and hepatopancreas (Li et al, 2015). Meanwhile, LvTRAF3 was also cloned and characterized from the shrimp L.vannamei; it had a transcript of 3,865 bp, with an ORF of 1,002 bp, and encoded a polypeptide of 333 amino acids, with a calculated molecular weight of 38.6 kDa (Li et al, 2020).TRAF3 functions as a regulator of innate immune response that involves many cellular processes.…”

Cloning and characterization of the LvCTL genes encoding C-type lectin from white-leg shrimp (Litopenaeus vannamei)

“…Tissue expression analysis showed LvCTL4 was distributed with high levels in the gills, intestine, epithelium and hepatopancreas (Li et al, 2015). Meanwhile, LvTRAF3 was also cloned and characterized from the shrimp L.vannamei; it had a transcript of 3,865 bp, with an ORF of 1,002 bp, and encoded a polypeptide of 333 amino acids, with a calculated molecular weight of 38.6 kDa (Li et al, 2020).TRAF3 functions as a regulator of innate immune response that involves many cellular processes When comparing to the L. vannamei genes, the predicted LvCTL3 sequence resulted in 97.28% similarity to the sequence of AGV68681 or 96.60% to XP_027212325, while LvCTL4 was 100% similar to the sequence of AKA64754 and XP_027228060. LvCTL3 and LvCTL4 were 60.69% (XP_042883674) and 80.43% (QEX50549) homologous to kuruma prawn (Penaeus japonicus), respectively.…”