A 3D Model of the Membrane Protein Complex Formed by the White Spot Syndrome Virus Structural Proteins

ABSTRACT The protein components of the white spot syndrome virus (WSSV) virion have been well established by proteomic methods, and at least 39 structural proteins are currently known. However, several details of the virus structure and assembly remain controversial, including the role of one of the major structural proteins, VP26. In this study, Triton X-100 was used in combination with various concentrations of NaCl to separate intact WSSV virions into distinct fractions such that each fraction contained envelope and tegument proteins, tegument and nucleocapsid proteins, or nucleocapsid proteins only. From the protein profiles and Western blotting results, VP26, VP36A, VP39A, and VP95 were all identified as tegument proteins distinct from the envelope proteins (VP19, VP28, VP31, VP36B, VP38A, VP51B, VP53A) and nucleocapsid proteins (VP664, VP51C, VP60B, VP15). We also found that VP15 dissociated from the nucleocapsid at high salt concentrations, even though DNA was still present. These results were confirmed by CsCl isopycnic centrifugation followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and liquid chromatography-nanoelectrospray ionization-tandem mass spectrometry, by a trypsin sensitivity assay, and by an immunogold assay. Finally, we propose an assembly process for the WSSV virion.

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Crystal Structures of Major Envelope Proteins VP26 and VP28 from White Spot Syndrome Virus Shed Light on Their Evolutionary Relationship

Journal of Virology ◽

10.1128/jvi.02505-06 ◽

2007 ◽

Vol 81 (12) ◽

pp. 6709-6717 ◽

Cited By ~ 64

Author(s):

Xuhua Tang ◽

Jinlu Wu ◽

J. Sivaraman ◽

Choy Leong Hew

Keyword(s):

Viral Infection ◽

White Spot Syndrome Virus ◽

Evolutionary Relationship ◽

Viral Proteins ◽

Envelope Proteins ◽

Viral Envelope ◽

White Spot ◽

Structural Proteins ◽

White Spot Syndrome ◽

Effective Transfer

ABSTRACT White spot syndrome virus (WSSV) is a virulent pathogen known to infect various crustaceans. It has bacilliform morphology with a tail-like appendage at one end. The envelope consists of four major proteins. Envelope structural proteins play a crucial role in viral infection and are believed to be the first molecules to interact with the host. Here, we report the localization and crystal structure of major envelope proteins VP26 and VP28 from WSSV at resolutions of 2.2 and 2.0 Å, respectively. These two proteins alone account for approximately 60% of the envelope, and their structures represent the first two structural envelope proteins of WSSV. Structural comparisons among VP26, VP28, and other viral proteins reveal an evolutionary relationship between WSSV envelope proteins and structural proteins from other viruses. Both proteins adopt β-barrel architecture with a protruding N-terminal region. We have investigated the localization of VP26 and VP28 using immunoelectron microscopy. This study suggests that VP26 and VP28 are located on the outer surface of the virus and are observed as a surface protrusion in the WSSV envelope, and this is the first convincing observation for VP26. Based on our studies combined with the literature, we speculate that the predicted N-terminal transmembrane region of VP26 and VP28 may anchor on the viral envelope membrane, making the core β-barrel protrude outside the envelope, possibly to interact with the host receptor or to fuse with the host cell membrane for effective transfer of the viral infection. Furthermore, it is tempting to extend this host interaction mode to other structural viral proteins of similar structures. Our finding has the potential to extend further toward drug and vaccine development against WSSV.

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Penaeidins are a novel family of antiviral effectors against WSSV in shrimp

10.1101/467571 ◽

2018 ◽

Author(s):

Bang Xiao ◽

Qihui Fu ◽

Shengwen Niu ◽

Haoyang Li ◽

Kai Lǚ ◽

...

Keyword(s):

Viral Infection ◽

Signaling Pathways ◽

Outer Surface ◽

Phagocytic Activity ◽

White Spot Syndrome Virus ◽

Antiviral Immunity ◽

White Spot ◽

Structural Proteins ◽

First Report ◽

White Spot Syndrome

AbstractPenaeidins are members of a family of key effectors with broad anti-bacterial activities in penaeid shrimp. However, the function of penaeidins in antiviral immunity is rarely reported and remains largely unknown. Herein, we uncovered that penaeidins are a novel family of antiviral effectors against white spot syndrome virus (WSSV). Firstly, RNAi in vivo mediated knockdown of each penaeidin from four identified penaeidins from Litopenaeus vannamei resulted in elevated viral loads and rendered shrimp more susceptible to WSSV, whilst the phenotype of survival rate in penaeidin-silenced shrimp can be rescued via the injection of recombinant penaeidin proteins. Moreover, pull-down assays demonstrated the conserved PEN domain of penaeidin was able to interact with WSSV structural proteins. Furthermore, we observed that colloidal gold-labeled penaeidins were located on the outer surface of the WSSV virion. By infection-blocking assay, we observed that hemocytes had lower viral infection rates in the group of WSSV preincubated with penaeidins than those of control group. Phagocytic activity analysis further showed that penaeidins were able to inhibit phagocytic activity of hemocytes against WSSV Taken together, these results suggest that penaeidins specifically binds to WSSV virion by interacting with its structural proteins, thus preventing viral infection that confers host against WSSV. In addition, dual-luciferase assay and EMSA assay demonstrated that penaeidins were regulated by Dorsal and Relish, two transcription factors of the canonical Toll and IMD pathway, respectively. To our best knowledge, this is the first report on uncovering the antiviral function of penaeidins in the innate immune system of shrimp.ImportancesWhite spot syndrome, caused by white spot syndrome virus (WSSV), is the most serious disease in shrimp aquaculture, which has long been a scourge of cultured shrimp industry. Herein, we provided some substantial evidences to indicate that penaeidins are a novel family of effectors with antiviral activity against WSSV in shrimp. Penaeidins such as BigPEN, PEN2 and PEN3 were able to interact with the outer surface of WSSV virion via binding to viral structural proteins, and thus preventing viral entry host cells. In addition, we demonstrated that the Toll and IMD signaling pathways can regulate the transcriptional expression of penaeidins, which may suggest an important role of the conserved innate signaling pathways in antiviral immunity. This is the first report of the antiviral mechanism of penaeidins in shrimp, which may provide some new insights into strategies to control WSSV infection in shrimp farms.

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Genomic and Proteomic Analysis of Thirty-Nine Structural Proteins of Shrimp White Spot Syndrome Virus

Journal of Virology ◽

10.1128/jvi.78.20.11360-11370.2004 ◽

2004 ◽

Vol 78 (20) ◽

pp. 11360-11370 ◽

Cited By ~ 174

Author(s):

Jyh-Ming Tsai ◽

Han-Ching Wang ◽

Jiann-Horng Leu ◽

He-Hsuan Hsiao ◽

Andrew H.-J. Wang ◽

...

Keyword(s):

White Spot Syndrome Virus ◽

Expression Patterns ◽

Sodium Dodecyl ◽

Peptide Sequence ◽

White Spot ◽

Structural Proteins ◽

Structural Protein ◽

Structural Genes ◽

Sds Page ◽

White Spot Syndrome

ABSTRACT White spot syndrome virus (WSSV) virions were purified from the hemolymph of experimentally infected crayfish Procambarus clarkii, and their proteins were separated by 8 to 18% gradient sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) to give a protein profile. The visible bands were then excised from the gel, and following trypsin digestion of the reduced and alkylated WSSV proteins in the bands, the peptide sequence of each fragment was determined by liquid chromatography-nano-electrospray ionization tandem mass spectrometry (LC-nanoESI-MS/MS) using a quadrupole/time-of-flight mass spectrometer. Comparison of the resulting peptide sequence data against the nonredundant database at the National Center for Biotechnology Information identified 33 WSSV structural genes, 20 of which are reported here for the first time. Since there were six other known WSSV structural proteins that could not be identified from the SDS-PAGE bands, there must therefore be a total of at least 39 (33 + 6) WSSV structural protein genes. Only 61.5% of the WSSV structural genes have a polyadenylation signal, and preliminary analysis by 3′ rapid amplification of cDNA ends suggested that some structural protein genes produced mRNA without a poly(A) tail. Microarray analysis showed that gene expression started at 2, 6, 8, 12, 18, 24, and 36 hpi for 7, 1, 4, 12, 9, 5, and 1 of the genes, respectively. Based on similarities in their time course expression patterns, a clustering algorithm was used to group the WSSV structural genes into four clusters. Genes that putatively had common or similar roles in the viral infection cycle tended to appear in the same cluster.

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Hijacking of Host Calreticulin is Required for the White Spot Syndrome Virus Replication Cycle.

Journal of Virology ◽

10.1128/jvi.01014-14 ◽

2014 ◽

pp. JVI.01014-14 ◽

Cited By ~ 5

Author(s):

Apiruck Watthanasurorot ◽

Enen Guo ◽

Sirinit Tharntada ◽

Chu-Fang Lo ◽

Kenneth Söderhäll ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

White Spot Syndrome Virus ◽

Replication Cycle ◽

Host Protein ◽

Host Cells ◽

White Spot ◽

Structural Proteins ◽

Dna Virus ◽

White Spot Syndrome ◽

Viral Structural Proteins

We have previously shown that multifunctional calreticulin (CRT), which resides in the endoplasmic reticulum (ER) and is involved in ER-associated protein processing, responds to infection with white spot syndrome virus (WSSV) by increasing mRNA and protein expression and by forming a complex with gC1qR and thereby delaying apoptosis. Here, we show that CRT can directly interact with WSSV structural proteins, including VP15 and VP28, during an early stage of virus infection. The binding of VP28 with CRT does not promote WSSV entry, and CRT-VP15 interaction was detected in the viral genome in virally infected host cells, and thus may have an effect on WSSV replication. Moreover, CRT was detected in the viral envelope of purified WSSV virions. CRT was also found to be of high importance for proper oligomerization of the viral structural proteins VP26 and VP28, and when CRT glycosylation was blocked with tunicamycin, a significant decrease in both viral replication and assembly was detected. Together, these findings suggest that CRT confers several advantages to WSSV from the initial steps of WSSV infection, to the assembly of virions. Therefore, CRT is required as a “vital factor” and is hijacked by WSSV for its replication cycle.ImportanceWhite spot syndrome virus (WSSV) is a double-stranded DNA virus and the cause of a serious disease in a wide range of crustaceans that often leads to high mortality rates. We have previously shown that the protein calreticulin (CRT), which resides in the endoplasmic reticulum (ER) of the cell, is important in the host response to the virus. In this report, we show that the virus uses this host protein to enter the cell and to make the host produce new viral structural proteins. Through its interaction with two viral proteins, the virus “hijacks” host calreticulin and uses it for its own needs. These findings provide new insight into the interaction between a large DNA virus and the host protein CRT, and may help in understanding the viral infection process in general.

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Identification of structural proteins from shrimp white spot syndrome virus (WSSV) by 2DE-MS

Proteins Structure Function and Bioinformatics ◽

10.1002/prot.10640 ◽

2004 ◽

Vol 55 (2) ◽

pp. 229-235 ◽

Cited By ~ 76

Author(s):

Xiaobo Zhang ◽

Canhua Huang ◽

Xuhua Tang ◽

Ying Zhuang ◽

Choy Leong Hew

Keyword(s):

White Spot Syndrome Virus ◽

White Spot ◽

Structural Proteins ◽

White Spot Syndrome

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Proteomic Analysis of the Major Envelope and Nucleocapsid Proteins of White Spot Syndrome Virus

Journal of Virology ◽

10.1128/jvi.01452-06 ◽

2006 ◽

Vol 80 (21) ◽

pp. 10615-10623 ◽

Cited By ~ 126

Author(s):

Xixian Xie ◽

Limei Xu ◽

Feng Yang

Keyword(s):

Posttranslational Modifications ◽

White Spot Syndrome Virus ◽

Procambarus Clarkii ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

White Spot ◽

Structural Proteins ◽

Nucleocapsid Proteins ◽

Flight Mass Spectrometry ◽

White Spot Syndrome

ABSTRACT White spot syndrome virus (WSSV) virions were purified from the tissues of infected Procambarus clarkii (crayfish) isolates. Pure WSSV preparations were subjected to Triton X-100 treatment to separate into the envelope and nucleocapsid fractions, which were subsequently separated by 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The major envelope and nucleocapsid proteins were identified by either matrix-assisted laser desorption ionization-time of flight mass spectrometry or defined antibody. A total of 30 structural proteins of WSSV were identified in this study; 22 of these were detected in the envelope fraction, 7 in the nucleocapsid fraction, and 1 in both the envelope and the nucleocapsid fractions. With the aid of specific antibodies, the localizations of eight proteins were further studied. The analysis of posttranslational modifications revealed that none of the WSSV structural proteins was glycosylated and that VP28 and VP19 were threonine phosphorylated. In addition, far-Western and coimmunoprecipitation experiments showed that VP28 interacted with both VP26 and VP24. In summary, the data obtained in this study should provide an important reference for future molecular studies of WSSV morphogenesis.

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