Full length articleeIF2α of Litopenaeus vannamei involved in shrimp immune response to WSSV infection
Introduction
Eukaryotic Initiation Factor 2 (eIF2) is an essential factor for initiation of protein synthesis. It is a heterotrimer that is composed of three subunits, namely α, β and γ. Its activity is regulated by a mechanism involving both guanine nucleotide exchange and phosphorylation. eIF2α is the main target for phosphorylation, which contains a serine at position 51. Therefore, the α subunit can be considered to be the regulatory subunit of the trimer, whose activity is regulated by phosphorylation [1]. The phosphorylation of eIF2α can result in a severe decline in protein synthesis, which is an important strategy in the cell's armory against stressful insults including viral infection, the accumulation of misfolded proteins, and starvation [2].
There are four well-characterized serine–threonine kinases, double-stranded RNA-activated protein kinase (PKR), PKR-like ER kinase (PERK), general control non-derepressible-2 (GCN2) and heme-regulated inhibitor (HRI), were reported to phosphorylate the eIF2α under different stresses [3], [4], [5], [6]. Among them, PKR can be activated by the dsRNA stimulation, and it's well known to play an important role in innate antiviral immunity [7]. And PERK is a sensor of endoplasmic reticulum (ER) stress. The endoplasmic reticulum is the place for the folding of proteins, and viral infection [8] or the over-expression of proteins [9] can lead to endoplasmic reticulum stress response. Unfolded protein response (UPR) is a cellular stress response related to the ER, and the studies indicated that virus infection could induce the ER stress and activate the UPR. Herpes simplex virus (HSV) infection induced phosphorylation of an endoplasmic reticulum (ER) resident kinase PERK [10], and African swine fever virus (ASFV) infection also induced transcription factor 6 signaling pathway of the UPR. The UPR was regulated by the virus to prevent the early apoptosis and facilitate viral replication [11].
Litopenaeus vannamei, as the main species of cultured shrimp, has been threatened by white spot syndrome virus (WSSV) [12]. Many studies on the innate immunity of shrimp have been carried out, and several immune-related genes in shrimp, such as hemocyanin [13], [14], lectin [15], [16] and antimicrobial peptide [17], [18] have been identified and characterized. Moreover, to understand the mechanisms involved in WSSV pathogenesis in shrimp, more and more studies had been focused on the signal transduction and some pathways have also been reported to be involved in the interaction between WSSV and shrimp [19], [20], [21], [22], [23]. Previous studies have indicated that there is a complex relationship between WSSV and UPR [24], [25], [26]. Activating transcription factor 4 of L. vannamei (LvATF4), a transcription factor of shrimp UPR, was activated under WSSV challenge [26], which may be regulated by the PERK/eIF2α.
To further understand the role of the UPR pathway between host and virus, the eIF2α of L. vannamei, designed as LveIF2α, was investigated under WSSV challenge in this study. The result suggested that LveIF2α was involved in WSSV infection.
Section snippets
Sequence analysis
Based on the analysis of L. vannamei (data not shown), the sequence of LveIF2α was obtained. The LveIF2α protein domains were predicted using the SMART program (http://smart.embl-heidelberg.de/). The sequences of eIF2α protein from other species in the database were searched and analyzed using the BLAST program (http://blast.ncbi.nlm.nih.gov/Blast.cgi). Multiple sequence alignment was performed using DNAMAN software. A neighbor-joining (NJ) phylogenic tree was constructed based on amino acid
Characterization of L. vannamei eIF2α
The full-length cDNA of LveIF2α is 2030 bp, including a 106 bp 5′ untranslated region (UTR) and a 928 bp 3′-UTR with a poly (A) tail. The open reading frame (ORF) of LveIF2α is 996 bp, which encodes a protein of 331 amino acids. The LveIF2α is highly homologous to other known eIF2α proteins, and also has S1_IF2_alpha domain at 13–88 aa and EIF_2_alpha domain at 129–242 aa, respectively, which are the characteristics of eIF2α. It also has a conserved protein synthesis regulator site at 51 aa,
Discussion
The WSSV is a serious pathogen in shrimp aquaculture, the studies of innate immune against virus have indicated several pathways were involved in WSSV infection [20], [22], [32]. The accumulation of unfolded or misfolded proteins in the lumen of the ER results in ER stress that triggers the UPR. Previous studies have shown WSSV can influence the transcription of the related genes in the UPR pathway [22], [24], [25], [26], which meant WSSV infection can cause the ER stress.
In this study,
Acknowledgments
This work was funded by the Projects under Major State Basic Research Development Program of China (973 Program) (No. 2012CB114403), the National High Technology Research and Development Program of China (863 Program) (No. 2012AA092205), the China Agriculture Research System-47, the National Natural Science Foundation of China (No. 31302209), and the Scientific Research Foundation of Third Institute of Oceanography, SOA (No. 2011018).
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