Modulation of the unfolded protein response by white spot syndrome virus via wsv406 targeting BiP to facilitate viral replication

Shihan Chen; Qiqi Zhong; Xuzheng Liao; Haiyang Wang; Bang Xiao; Jianguo He; Chaozheng Li

doi:10.1016/j.virs.2024.10.005

December 2024

. doi: 10.1016/j.virs.2024.10.005

Citation: Shihan Chen, Qiqi Zhong, Xuzheng Liao, Haiyang Wang, Bang Xiao, Jianguo He, Chaozheng Li. Modulation of the unfolded protein response by white spot syndrome virus via wsv406 targeting BiP to facilitate viral replication .VIROLOGICA SINICA, 2024, 39(6) : 938-950. http://dx.doi.org/10.1016/j.virs.2024.10.005

白斑综合征病毒通过wsv406靶向BiP调控未折叠蛋白反应促进病毒复制

陈史瀚 ^a,b ,
钟琪琪 ^a,b ,
廖栩峥 ^a,b ,
王海阳 ^a,b ,
肖邦 ^a,b ,
何建国 ^a,b,c ,
李朝政 ^a,b,c,,

cstr: 32224.14.j.virs.2024.10.005

a. 中山大学海洋科学学院, 水产动物疫病防控与健康养殖全国重点实验室/南方海洋科学与工程广东省实验室(珠海), 广州 510275;
b. 广东省海洋资源与近岸工程重点实验室/广东省水生经济动物良种繁育重点实验室, 中山大学, 广州 510275;
c. 中国-东盟海水养殖技术"一带一路"联合实验室, 中山大学, 广州 510275

通讯作者： 李朝政, lichaozh@mail2.sysu.edu.cn
收稿日期： 2024-06-14
录用日期： 2024-10-24

摘要

疾病的爆发通常与环境压力有关，这可能导致内质网（ER）压力并随后触发未折叠蛋白反应（UPR）。对虾养殖中最严重的病原体——白斑综合征病毒（WSSV）的复制已被证明依赖于UPR信号通路，但其详细机制尚不清楚。本文中，研究发现WSSV通过病毒蛋白wsv406劫持UPR通路来增强其复制。分析显示，在感染WSSV对虾的血细胞和鳃中，wsv406显著上调。质谱分析表明wsv406能与凡纳滨对虾（Litopenaeus vannamei）免疫球蛋白重链结合蛋白（BiP）相互作用。进一步研究发现，wsv406能结合LvBiP的多个结构域，抑制其ATP酶活性，但不破坏其与UPR压力受体的结合。沉默wsv406或LvBiP导致WSSV复制减少，并提高对虾的存活率。此外，在WSSV感染期间沉默wsv406，eIF2α的磷酸化和ATF6的核易位显著降低，提示wsv406激活了PRKR样ER激酶(PERK)-真核翻译起始因子2α (eIF2α)和激活转录因子6 （ATF6）途径。这种激活促进了WSSV基因的转录和病毒的复制。综上所述，这些发现表明wsv406通过靶向LvBiP调控宿主UPR，进而通过PERK-eIF2α和ATF6途径增强WSSV的复制。这揭示了WSSV与宿主细胞机制之间的相互作用，为开发治疗性干预措施以控制对虾养殖中WSSV的爆发提供了潜在的靶标。
- 对虾
- , 白斑综合征病毒
- , 未折叠蛋白反应
- , wsv406
- , 结合蛋白（BiP）

Modulation of the unfolded protein response by white spot syndrome virus via wsv406 targeting BiP to facilitate viral replication

Shihan Chen ^a,b ,
Qiqi Zhong ^a,b ,
Xuzheng Liao ^a,b ,
Haiyang Wang ^a,b ,
Bang Xiao ^a,b ,
Jianguo He ^a,b,c ,
Chaozheng Li ^a,b,c,,

a. School of Marine Sciences, Sun Yat-sen University, State Key Laboratory of Biocontrol/ Southern Marine Science and Engineering Guangdong Laboratory (Zhuhai), Guangzhou, 510275, China;
b. Guangdong Provincial Key Laboratory of Marine Resources and Coastal Engineering/ Guangdong Provincial Key Laboratory for Aquatic Economic Animals, Sun Yat-sen University, Guangzhou, 510275, China;
c. China-ASEAN Belt and Road Joint Laboratory on Marine Aquaculture Technology, Sun Yat-sen University, Guangzhou, 510275, China

Corresponding author: Chaozheng Li, lichaozh@mail2.sysu.edu.cn
Received Date: 14 June 2024
Accepted Date: 24 October 2024

Abstract

Outbreaks of diseases are often linked to environmental stress, which can lead to endoplasmic reticulum (ER) stress and subsequently trigger the unfolded protein response (UPR). The replication of the white spot syndrome virus (WSSV), the most serious pathogen in shrimp aquaculture, has been shown to rely on the UPR signaling pathway, although the detailed mechanisms remain poorly understood. In this study, we discovered that WSSV enhances its replication by hijacking the UPR pathway via the viral protein wsv406. Our analysis revealed a significant upregulation of wsv406 in the hemocytes and gills of infected shrimp. Mass spectrometry analysis identified that wsv406 interacts specifically with the immunoglobulin heavy-chain-binding protein (BiP) in shrimp Litopenaeus vannamei. Further examination revealed that wsv406 binds to multiple domains of LvBiP, inhibiting its ATPase activity without disrupting its binding to UPR stress receptors. Silencing either wsv406 or LvBiP resulted in a reduction in WSSV replication and improved shrimp survival rates. Further, wsv406 activation of the PRKR-like ER kinase (PERK)-eukaryotic translation initiation factor 2α (eIF2α) and activating transcription factor 6 (ATF6) pathways was demonstrated by a decrease in the phosphorylation of eIF2α and the nuclear translocation of ATF6 when wsv406 was silenced during WSSV infection. This activation facilitated the transcription of WSSV genes, promoting viral replication. In summary, these findings reveal that wsv406 manipulates the host UPR by targeting LvBiP, thereby enhancing WSSV replication through the PERK-eIF2α and ATF6 pathways. These insights into the interaction between WSSV and host cellular machinery offer potential targets for developing therapeutic interventions to control WSSV outbreaks in shrimp aquaculture.
- Shrimp
- , White spot syndrome virus
- , Unfolded protein response
- , wsv406
- , Binding protein (BiP)

References
1. Ai, H. S., Liao, J. X., Huang, X. D., Yin, Z. X., Weng, S. P., Zhao, Z. Y., Li, S. D., Yu, X. Q., He, J. G., 2009. A novel prophenoloxidase 2 exists in shrimp hemocytes. Dev. Comp. Immunol. 33, 59-68.
2. Arulmoorthy, M. P., Anandajothi, E., Vasudevan, S., Suresh, E., 2020. Major viral diseases in culturable penaeid shrimps: a review. Aquacult. Int. 28, 1939-1967.
3. Bertolotti, A., Zhang, Y. H., Hendershot, L. M., Harding, H. P., Ron, D., 2000. Dynamic interaction of BiP and ER stress transducers in the unfolded-protein response. Nat. Cell Biol. 2, 326-332.
4. Chen, Y. H., Zhao, L., Pang, L. R., Li, X. Y., Weng, S. P., He, J. G., 2012. Identification and characterization of Inositol-requiring enzyme-1 and X-box binding protein 1, two proteins involved in the unfolded protein response of Litopenaeus vannamei. Dev. Comp. Immunol. 38, 66-77.
5. Chen, Y. H., Yuan, F. H., Bi, H. T., Zhang, Z. Z., Yue, H. T., Yuan, K., Chen, Y. G., Weng, S. P., He, J. G., 2016. Transcriptome analysis of the unfolded protein response in hemocytes of Litopenaeus vannamei. Fish Shellfish Immunol. 54, 153-163.
6. Chen, T. H., Chiang, Y. H., Hou, J. N., Cheng, C. C., Sofiyatun, E., Chiu, C. H., Chen, W. J., 2017. XBP1-mediated BiP/GRP78 upregulation copes with oxidative stress in mosquito cells during dengue 2 virus infection. Biomed Res. Int. 2017, 3519158.
7. Elfiky, A. A., 2020. SARS-CoV-2 spike-heat shock protein A5 (GRP78) recognition may be related to the immersed human coronaviruses. Front. Pharmacol. 11, 577467.
8. Galindo, I., Hernaez, B., Munoz-Moreno, R., Cuesta-Geijo, M. A., Dalmau-Mena, I., Alonso, C., 2012. The ATF6 branch of unfolded protein response and apoptosis are activated to promote African swine fever virus infection. Cell Death Dis. 3, e341.
9. Gething, M. J., 1999. Role and regulation of the ER chaperone BiP. Semin. Cell Dev. Biol. 10, 465-472.
10. He, F., Fenner, B. J., Godwin, A. K., Kwang, J., 2006. White spot syndrome virus open reading frame 222 encodes a viral E3 ligase and mediates degradation of a host tumor suppressor via ubiquitination. J. Virol. 80, 3884-3892.
11. Ibrahim, I. M., Abdelmalek, D. H., Elshahat, M. E., Elfiky, A. A., 2020. COVID-19 spike-host cell receptor GRP78 binding site prediction. J. Infect. 80, 554-562.
12. Isler, J. A., Skalet, A. H., Alwine, J. C., 2005. Human cytomegalovirus infection activates and regulates the unfolded protein response. J. Virol. 79, 6890-6899.
13. Janewanthanakul, S., Supungul, P., Tang, S., Tassanakajon, A., 2020. Heat shock protein 70 from Litopenaeus vannamei (LvHSP70) is involved in the innate immune response against white spot syndrome virus (WSSV) infection. Dev. Comp. Immunol. 102, 103476.
14. Jiang, S. G., Qiu, L. H., Zhou, F. L., Huang, J. H., Guo, Y. H., Yang, K., 2007. Molecular cloning and expression analysis of a heat shock protein (Hsp90) gene from black tiger shrimp (Penaeus monodon). Mol. Biol. Rep. 36, 127-134.
15. Jiang, Z. H., Zhang, K., Li, Z. L., Li, Z. G., Yang, M., Jin, X. J., Cao, Q., Wang, X. T., Yue, N., Li, D. W. et al., 2020. The Barley stripe mosaic virus γb protein promotes viral cell-to-cell movement by enhancing ATPase-mediated assembly of ribonucleoprotein movement complexes. PLoS Pathog. 16, e1008709.
16. Li, X. Y., Pang, L. R., Chen, Y. G., Weng, S. P., Yue, H. T., Zhang, Z. Z., Chen, Y. H., He, J. G., 2013. Activating transcription factor 4 and X box binding protein 1 of Litopenaeus vannamei transcriptional regulated white spot syndrome virus genes wsv023 and wsv083. PLoS One 8, e62603.
17. Li, X. Y., Yue, H. T., Zhang, Z. Z., Bi, H. T., Chen, Y. G., Weng, S. P., Chan, S., He, J. G., Chen, Y. H., 2014. An activating transcription factor of Litopenaeus vannamei involved in WSSV genes wsv059 and wsv166 regulation. Fish Shellfish Immunol. 41, 147-155.
18. Li, C. Y., Wang, Y. J., Huang, S. W., Cheng, C. S., Wang, H. C., 2016. Replication of the shrimp virus WSSV depends on glutamate-driven anaplerosis. PLoS One 11, e0146902.
19. Li, H. Y., Wang, S., Chen, Y. G., Lǔ, K., Yin, B., Li, S. D., He, J. G., Li, C. Z., 2017. Identification of two p53 isoforms from Litopenaeus vannamei and their interaction with NF-κB to induce distinct immune response. Sci. Rep. 7, 45821.
20. Li, H. Y., Yin, B., Wang, S., Fu, Q. H., Xiao, B., Lǔ, K., He, J. G., Li, C. Z., 2018. RNAi screening identifies a new Toll from shrimp Litopenaeus vannamei that restricts WSSV infection through activating Dorsal to induce antimicrobial peptides. PLoS Pathog. 14, e1007109.
21. Li, C. Z., Weng, S. P., He, J. G., 2019. WSSV-host interaction: Host response and immune evasion. Fish Shellfish Immunol. 84, 558-571.
22. Luan, W., Li, F. H., Zhang, J. Q., Wang, B., Xiang, J. H., 2009. Cloning and expression of glucose regulated protein 78 (GRP78) in Fenneropenaeus chinensis. Mol. Biol. Rep. 36, 289-298.
23. Luana, W., Li, F. H., Wang, B., Zhang, X. J., Liu, Y. C., Xiang, J. H., 2007. Molecular characteristics and expression analysis of calreticulin in Chinese shrimp Fenneropenaeus chinensis. Comp. Biochem. Phys. B Biochem. Mol. Biol. 147, 482-491.
24. Millard, R. S., Ellis, R. P., Bateman, K. S., Bickley, L. K., Tyler, C. R., van Aerle, R., Santos, E. M., 2021. How do abiotic environmental conditions influence shrimp susceptibility to disease? A critical analysis focussed on White Spot Disease. J. Invertebr. Pathol. 186, 107369.
25. Paskevicius, T., Farraj, R. A., Michalak, M., Agellon, L. B., 2023. Calnexin, more than just a molecular chaperone. Cells 12, 403.
26. Pena, J., Harris, E., 2011. Dengue virus modulates the unfolded protein response in a time-dependent manner. J. Biol. Chem. 286, 14226-14236.
27. Schroder, M., Kaufman, R. J., 2005. The mammalian unfolded protein response. Annu. Rev. Biochem. 74, 739-789.
28. Shen, J. S., Chen, X., Hendershot, L., Prywes, R., 2002. ER stress regulation of ATF6 localization by dissociation of BiP/GRP78 binding and unmasking of golgi localization signals. Dev. Cell 3, 99-111.
29. Shin, W. J., Ha, D. P., Machida, K., Lee, A. S., 2022. The stress-inducible ER chaperone GRP78/BiP is upregulated during SARS-CoV-2 infection and acts as a pro-viral protein. Nat. Commun. 13, 6551.
30. Siddique, M. A., Haque, M. I., Sanyal, S. K., Hossain, A., Nandi, S. P., Alam, A., Sultana, M., Hasan, M., Hossain, M. A., 2018. Circulatory white spot syndrome virus in South-West region of Bangladesh from 2014 to 2017: molecular characterization and genetic variation. AMB Express 8, 25.
31. Stentiford, G. D., Neil, D. M., Peeler, E. J., Shields, J. D., Small, H. J., Flegel, T. W., Vlak, J. M., Jones, B., Morado, F., Moss, S. et al., 2012. Disease will limit future food supply from the global crustacean fishery and aquaculture sectors. J. Invertebr. Pathol. 110, 141-157.
32. Tassanakajon, A., Amparyup, P., Somboonwiwat, K., Supungul, P., 2011. Cationic antimicrobial peptides in penaeid shrimp. Mar. Biotechnol. 13, 639-657.
33. Umareddy, I., Pluquet, O., Wang, Q. Y., Vasudevan, S. G., Chevet, E., Gu, F., 2007. Dengue virus serotype infection specifies the activation of the unfolded protein response. Virol. J. 4, 91.
34. Van Etten, J., 2009. Lesser known large dsDNA viruses. Preface. Curr. Top. Microbiol. 328, v-vii.
35. Walter, P., Ron, D., 2011. The unfolded protein response: from stress pathway to homeostatic regulation. Science 334, 1081-1086.
36. Wang, M., Kaufman, R. J., 2014. The impact of the endoplasmic reticulum protein-folding environment on cancer development. Nat. Rev. Cancer 14, 581-597.
37. Wang, S., Li, H. Y., Zhu, P., Fu, Q. H., Yin, B., Li, Q. Y., Chen, R. J., Jiang, X. W., Weng, S. P., He, J.G. et al., 2021. MAPKKK15 gene from shrimp Litopenaeus vannamei is transcribed in larva development stages and contributes to WSSV pathogenesis. Aquaculture 534, 736324.
38. Wang, C. Q., Wei, M. H., Wu, G. C., He, L. X., Zhu, J. H., Aweya, J. J., Chen, X. L., Zhao, Y. Z., Zhang, Y. L., Yao, D. F., 2022. Proteomics analysis reveals a critical role for the WSSV immediate-early protein IE1 in modulating the host prophenoloxidase system. Virulence 13, 936-948.
39. Xiao, B., Fu, Q. H., Niu, S. W., Zhu, P., He, J. G., Li, C. Z., 2020. Penaeidins restrict white spot syndrome virus infection by antagonizing the envelope proteins to block viral entry. Emerg. Microbes Infec. 9, 390-412.
40. Xu, J. X., Ruan, L. W., Shi, H., 2014. eIF2α of Litopenaeus vannamei involved in shrimp immune response to WSSV infection. Fish Shellfish Immunol. 40, 609-615.
41. Yan, D. C., Dong, S. L., Huang, J., Yu, X. M., Feng, M. Y., Liu, X. Y., 2004. White spot syndrome virus (WSSV) detected by PCR in rotifers and rotifer resting eggs from shrimp pond sediments. Dis. Aquat. Organ. 59, 69-73.
42. Yu, C. Y., Hsu, Y. W., Liao, C. L., Lin, Y. L., 2006. Flavivirus infection activates the XBP1 pathway of the unfolded protein response to cope with endoplasmic reticulum stress. J. Virol. 80, 11868-11880.
43. Yuan, F. H., Chen, Y. G., Zhang, Z. Z., Yue, H. T., Bi, H. T., Yuan, K., Weng, S. P., He, J. G., Chen, Y. H., 2016. Down-regulation apoptosis signal-regulating kinase 1 gene reduced the Litopenaeus vannamei hemocyte apoptosis in WSSV infection. Fish Shellfish Immunol. 50, 109-116.
44. Yuan, K., He, H. H., Zhang, C. Z., Li, X. Y., Weng, S. P., He, J. G., Chen, Y. H., 2017. Litopenaeus vannamei activating transcription factor 6 alpha gene involvement in ER-stress response and white spot symptom virus infection. Fish Shellfish Immunol. 70, 129-139.
45. Yuan, Z. Z., Chen, M., Wang, J. T., Li, Z. Y., Geng, X. Y., Sun, J. S., 2018. Identification of Litopenaeus vannamei BiP as a novel cellular attachment protein for white spot syndrome virus by using a biotinylation based affinity chromatography method. Fish Shellfish Immunol. 79, 130-139.
Proportional views
Electronic Supplementary Material

10.1016j.virs.2024.10.005-ESM.docx