DOI QR코드

DOI QR Code

Molecular characterization of interferon regulatory factors (IRFs) 8 of red sea bream (Pagrus major) and expression analysis after RSIV infection

  • Won-Sik Woo (Department of Marine Biology & Aquaculture, College of Marine Science, Gyeongsang National University) ;
  • Gyoungsik Kang (Department of Marine Biology & Aquaculture, College of Marine Science, Gyeongsang National University) ;
  • Kyung-Ho Kim (Department of Marine Biology & Aquaculture, College of Marine Science, Gyeongsang National University) ;
  • Ha-Jeong Son (Department of Marine Biology & Aquaculture, College of Marine Science, Gyeongsang National University) ;
  • Min-Young Sohn (Department of Marine Biology & Aquaculture, College of Marine Science, Gyeongsang National University) ;
  • Ju-Won Kim (Biotechnology Research Division, National Institute of Fisheries Science) ;
  • Hee Jeong Kong (Biotechnology Research Division, National Institute of Fisheries Science) ;
  • Young-Ok Kim (Biotechnology Research Division, National Institute of Fisheries Science) ;
  • Chan-Il Park (Department of Marine Biology & Aquaculture, College of Marine Science, Gyeongsang National University)
  • Received : 2023.04.27
  • Accepted : 2023.05.11
  • Published : 2023.06.30

Abstract

Interferon regulatory factors (IRFs) exhibit antiviral and antibacterial properties. In this study, we cloned the IRF8 genes of red sea bream (Pagrus major) and analyzed their expression following red sea bream iridovirus (RSIV) infection. Multi-sequence analysis revealed conserved features, such as the IRF-association domain, nuclear localization signal, and DNA-binding domain. Phylogenetic analysis demonstrated that PmIRF8 clustered with IRF8 from other fish species. Quantitative real-time PCR was employed to examine IRF8 expression in normal tissues, revealing high expression levels in the gill (26.7-fold), spleen (25.7-fold) and brain (22.6-fold). This finding aligns with previous research, which has established that IRF8 is associated with immune cell development and is highly expressed in lymphomyeloid tissues. Additionally, temporal expression of PmIRF8 following RSIV infection demonstrated an initial low expression in the gills, followed by an upregulation at 36 hours post-infection (hpi). A similar expression pattern was observed in the spleen. The liver showed variable expression, while the kidney exhibited an increase at 6 hpi, followed by another upregulation at 3 days post-infection. These results suggest that PmIRF8 plays a role in regulation and suppression during the immune response. These findings are corroborated by previous studies on other fish species. Overall, this investigation presents the characterization of IRF genes in red sea bream and provides foundational data for comparative analysis of IRF genes.

Keywords

Acknowledgement

This research was supported by the National Institute of Fisheries Science, Ministry of Oceans and Fisheries, Korea (R2023024)

References

  1. Alter-Koltunoff, M., Goren, S., Nousbeck, J., Feng, C. G., Sher, A., Ozato, K., Azriel, A., & Levi, B.-Z. Innate immunity to intraphagosomal pathogens is mediated by interferon regulatory factor 8 (IRF-8) that stimulates the expression of macrophage-specific Nramp1 through antagonizing repression by c-Myc. Journal of Biological Chemistry, 283(5), 2724-2733. https://doi.org/10.1074/jbc.M707704200.2008.
  2. Andrilenas, K. K., Ramlall, V., Kurland, J., Leung, B., Harbaugh, A. G., & Siggers, T. DNA-binding landscape of IRF3, IRF5 and IRF7 dimers: implications for dimer-specific gene regulation. Nucleic acids research, 46(5), 2509-2520. https://doi.org/10.1093/nar/gky002 2018.
  3. Antonczyk, A., Krist, B., Sajek, M., Michalska, A., Piaszyk-Borychowska, A., Plens-Galaska, M., Wesoly, J., & Bluyssen, H. A. R. Direct Inhibition of IRF-Dependent Transcriptional Regulatory Mechanisms Associated With Disease. Front Immunol, 10, 1176. https://doi.org/10.3389/fimmu.2019.01176 2019.
  4. Bathige, S., Whang, I., Umasuthan, N., Lim, B.-S., Park, M.-A., Kim, E., Park, H.-C., & Lee, J. Interferon regulatory factors 4 and 8 in rock bream, Oplegnathus fasciatus: structural and expressional evidence for their antimicrobial role in teleosts. Fish & Shellfish Immunology, 33(4), 857-871. 2012. https://doi.org/10.1016/j.fsi.2012.07.017
  5. Chang, K., Han, K., Qiu, W., Hu, Z., Chen, X., Chen, X., Xie, X., Wang, S., Hu, C., & Mao, H. Grass carp (Ctenopharyngodon idella) interferon regulatory factor 8 down-regulates interferon1 expression via interaction with interferon regulatory factor 2 in vitro. Molecular Immunology, 137, 202-211. https://www.sciencedirect.com/science/article/pii/S0161589021001371?via%3Dihub 2021. 1001371?via%3Dihub
  6. Chen, X., Hu, G., Dong, X., Liu, Q., & Zhang, S. Molecular cloning and expression analysis of interferon regulatory factor 8 (IRF8) in turbot, Scophthalmus maximus. Veterinary immunology and immunopathology, 149(1-2), 143-150. 2012. https://doi.org/10.1016/j.vetimm.2012.06.008
  7. Das, A., Chauhan, K. S., Kumar, H., & Tailor, P. Mutation in Irf8 Gene (Irf8R294C) Impairs Type I IFN-Mediated Antiviral Immune Response by Murine pDCs [Original Research]. Frontiers in immunology, 12. https://doi.org/10.3389/fimmu.2021.758190 2021.
  8. Gabriele, L., & Ozato, K. The role of the interferon regulatory factor (IRF) family in dendritic cell development and function. Cytokine & growth factor re- views, 18(5-6), 503-510. 2007. https://doi.org/10.1016/j.cytogfr.2007.06.008
  9. Girisha, S., Puneeth, T., Nithin, M., Kumar, B. N., Ajay, S., Vinay, T., Suresh, T., Venugopal, M., & Ramesh, K. Red sea bream iridovirus disease (RSIVD) outbreak in Asian seabass (Lates calcarifer) cultured in open estuarine cages along the west coast of India: First report. Aquaculture, 520, 734712. https://doi.org/10.1016/j.aquaculture.2019.734712 2020.
  10. Guan, Y., Chen, X., Luo, T., Ao, J., Ai, C., & Chen, X. Molecular characterization of the interferon regulatory factor (IRF) family and functional analysis of IRF11 in the large yellow croaker (Larimichthys crocea). Fish & Shellfish Immunology, 107, 218-229. https://doi.org/10.1016/j.fsi.2020.10.001. 2020.
  11. Haque, M. N., Nam, S.-E., Shin, Y. K., & Rhee, J.-S. The dinoflagellate Alexandrium affine acutely induces significant modulations on innate immunity, hepatic function, and antioxidant defense system in the gill and liver tissues of red seabream. Aquatic Toxicology, 240, 105985. https://doi.org/10.1016/j.aquatox.2021.105985 2021.
  12. Holland, J. W., Karim, A., Wang, T., Alnabulsi, A., Scott, J., Collet, B., Mughal, M. S., Secombes, C. J., & Bird, S. Molecular cloning and characterization of interferon regulatory factors 4 and 8 (IRF-4 and IRF-8) in rainbow trout, Oncorhynchus mykiss. Fish & Shellfish Immunology, 29(1), 157-166. 2010. https://doi.org/10.1016/j.fsi.2010.03.001
  13. Holtschke, T., Lohler, J., Kanno, Y., Fehr, T., Giese, N., Rosenbauer, F., Lou, J., Knobeloch, K.-P., Gabriele, L., & Waring, J. F. Immunodeficiency and chronic myelogenous leukemia-like syndrome in mice with a targeted mutation of the ICSBP gene. Cell, 87(2), 307-317. 1996. https://doi.org/10.1016/S0092-8674(00)81348-3
  14. Hu, G., Chen, X., Gong, Q., Liu, Q., Zhang, S., & Dong, X. Structural and expression studies of interferon regulatory factor 8 in Japanese flounder, Paralichthys olivaceus. Fish & Shellfish Immunology, 35(3), 1016-1024. 2013. https://doi.org/10.1016/j.fsi.2013.06.016
  15. Inouye, K., Yamano, K., Maeno, Y., Nakajima, K., Matsuoka, M., Wada, Y., & Sorimachi, M. Iridovirus infection of cultured red sea bream, Pagrus major. Fish Pathology, 27(1), 19-27. 1992. https://doi.org/10.3147/jsfp.27.19
  16. Joo, M.-S., Choi, K.-M., Kang, G., Woo, W.-S., Kim, K.-H., Sohn, M.-Y., Son, H.-J., Lee, J.-H., & Park, C.-I. Molecular characteristics, expression analysis and antiviral activity against RSIV of red sea bream IRF3 and IRF7. Aquaculture Reports, 27, 101415. https://doi.org/10.1016/j.aqrep.2022.101415 2022.
  17. Kim, K.-H., Choi, K.-M., Joo, M.-S., Kang, G., Woo, W.-S., Sohn, M.-Y., Son, H.-J., Kwon, M.-G., Kim, J.-O., & Kim, D.-H. Red Sea Bream Iridovirus (RSIV) Kinetics in Rock Bream (Oplegnathus fasciatus) at Various Fish-Rearing Seawater Temperatures. Animals, 12(15), 1978. 2022a.
  18. Kim, K.-H., Choi, K.-M., Kang, G., Woo, W.-S., Sohn, M.-Y., Son, H.-J., Yun, D., Kim, D.-H., & Park, C.-I. Development and Validation of a Quantitative Polymerase Chain Reaction Assay for the Detection of Red Sea Bream Iridovirus. Fishes, 7(5), 236. 2022b.
  19. Kim, K.-H., Joo, M.-S., Kang, G., Woo, W.-S., Sohn, M.-Y., Son, H.-J., & Park, C.-I. Characterization of Red Sea Bream (Pagrus major) Interferon Regulatory Factor 5 and 6 Genes and Their Expression in Response to RSIV Infection. Fishes, 8(2), 114. https://doi.org/10.3390/fishes8020114 2023.
  20. Kim, K., Hwang, S., Cho, M., Jung, S., Kim, Y., & Jeong, H. A natural infection by the red sea bream iridovirus-type Megalocytivirus in the golden mandarin fish Siniperca scherzeri. Journal of fish diseases, 41(8), 1229-1233. https://doi.org/10.1111/jfd.128152018.
  21. Kwon, W. J., Yoon, M. J., Jin, J. W., Kim, K. I., Kim, Y. C., Hong, S., Jeong, J. B., & Do Jeong, H. Development and characterization of megalocytivirus persistently-infected cell cultures for high yield of virus. Tissue and Cell, 66, 101387. 2020.
  22. Laghari, Z. A., Li, L., Chen, S. N., Huo, H. J., Huang, B., Zhou, Y., & Nie, P. Composition and transcription of all interferon regulatory factors (IRFs), IRF1‒11 in a perciform fish, the mandarin fish, Siniperca chuatsi. Developmental & Comparative Immunology, 81, 127-140. 2018. https://doi.org/10.1016/j.dci.2017.11.015
  23. Langlais, D., Barreiro, L. B., & Gros, P. The macrophage IRF8/IRF1 regulome is required for protection against infections and is associated with chronic inflammation. Journal of Experimental Medicine, 213(4), 585-603. 2016. https://doi.org/10.1084/jem.20151764
  24. Levi, B.-Z., Hashmueli, S., Gleit-Kielmanowicz, M., Azriel, A., & Meraro, D. ICSBP/IRF-8 transactivation: a tale of protein-protein interaction. Journal of interferon & cytokine research, 22(1), 153-160. 2002. https://doi.org/10.1089/107999002753452764
  25. Li, L., Jin, H., Xu, J., Shi, Y., & Wen, Z. Irf8 regulates macrophage versus neutrophil fate during zebrafish primitive myelopoiesis. Blood, The Journal of the American Society of Hematology, 117(4), 1359-1369. 2011. https://doi.org/10.1182/blood-2010-06-290700
  26. Livak, K. J., & Schmittgen, T. D. Analysis of relative gene expression data using real-time quantitative PCR and the 2- ΔΔCT method. Methods, 25(4), 402-408. 2001. https://doi.org/10.1006/meth.2001.1262
  27. Lopez-Porras, A., Morales, J. A., Alvarado, G., Koda, S. A., Camus, A., Subramaniam, K., Waltzek, T. B., & Soto, E. Red seabream iridovirus associated with cultured Florida pompano Trachinotus carolinus mortality in Central America. Diseases of aquatic organisms, 130(2), 109-115. https://doi.org/10.3354/dao03267. 2018.
  28. Lu, R. Interferon regulatory factor 4 and 8 in B-cell development. Trends Immunol, 29(10), 487-492. https://doi.org/10.1016/j.it.2008.07.006 2008.
  29. Medzhitov, R., & Janeway Jr, C. The Toll receptor family and microbial recognition. Trends in microbiology, 8(10), 452-456. 2000. https://doi.org/10.1016/S0966-842X(00)01845-X
  30. Nehyba, J., Hrdlickova, R., Burnside, J., & Bose Jr, H. R. A novel interferon regulatory factor (IRF), IRF-10, has a unique role in immune defense and is induced by the v-Rel oncoprotein. Molecular and cellular biology, 22(11), 3942-3957. 2002. https://doi.org/10.1128/MCB.22.11.3942-3957.2002
  31. Paun, A., & Pitha, P. The IRF family, revisited. Biochimie, 89(6-7), 744-753. 2007. https://doi.org/10.1016/j.biochi.2007.01.014
  32. Periyasamy, T., & Lu, M.-W. Molecular characterization and expression of malabar grouper (Epinephelus malabaricus) IRF8 gene response to immuno-stimulants and nervous necrosis virus. pathways, 8, 16. 2018.
  33. Puneeth, T., Baliga, P., Girisha, S., Shekar, M., Nithin, M., Suresh, T., & Kumar, B. N. Complete genome analysis of a red seabream iridovirus (RSIV) isolated from Asian seabass (Lates calcarifer) in India. Virus research, 291, 198199. https://doi.org/10.1016/j.virusres.2020.198199 2021.
  34. Rosenbauer, F., Waring, J. F., Foerster, J., Wietstruk, M., Philipp, D., & Horak, I. Interferon consensus sequence binding protein and interferon regulatory factor-4/Pip form a complex that represses the expression of the interferon-stimulated gene-15 in macrophages. Blood, The Journal of the American Society of Hematology, 94(12), 4274-4281. 1999. https://doi.org/10.1182/blood.V94.12.4274.424k05_4274_4281
  35. Savitsky, D., Tamura, T., Yanai, H., & Taniguchi, T. Regulation of immunity and oncogenesis by the IRF transcription factor family. Cancer immunology, immunotherapy, 59(4), 489-510. https://doi.org/10.1007/s00262-009-0804-6. 2010.
  36. Seo, J. S., Haque, M. N., Nam, S.-E., Kim, B.-M., & Rhee, J.-S. Inorganic nitrogen compounds reduce immunity and induce oxidative stress in red seabream. Fish & Shellfish Immunology, 104, 237-244. https://doi.org/10.1016/j.fsi.2020.05.072 2020.
  37. Sharf, R., Azriel, A., Lejbkowicz, F., Winograd, S. S., Ehrlich, R., & Levi, B.-Z. Functional Domain Analysis of Interferon Consensus Sequence Binding Protein (ICSBP) and Its Association with Interferon Regulatory Factors∗. Journal of Biological Chemistry, 270(22), 13063-13069. 1995. https://doi.org/10.1074/jbc.270.22.13063
  38. Shu, C., Sun, Y., & Xu, T. Molecular characterization of three IRF1 subfamily members reveals evolutionary significance of IRF11 in miiuy croaker. Developmental & Comparative Immunology, 53(2), 385-391. https://doi.org/10.1016/j.dci.2015.07.009.2015.
  39. Suzuki, Y., Yasuike, M., Kondo, H., Aoki, T., & Hirono, I. Molecular cloning and expression analysis of interferon regulatory factor 10 (IRF10) in Japanese flounder, Paralichthys olivaceus. Fish & Shellfish Immunology, 30(1), 67-76. https://doi.org/10.1016/j.fsi.2010.09.010. 2011.
  40. Tripathi, A., Singh Rawat, B., Addya, S., Surjit, M., Tailor, P., Vrati, S., & Banerjee, A. Lack of Interferon (IFN) Regulatory Factor 8 Associated with Restricted IFN-γ Response Augmented Japanese Encephalitis Virus Replication in the Mouse Brain. J Virol, 95 (21), e0040621. https://doi.org/10.1128/jvi.00406-21 2021.
  41. Wang, H., & Morse, H. C., 3rd. IRF8 regulates myeloid and B lymphoid lineage diversification. Immunologic research, 43(1-3), 109-117. https://doi.org/10.1007/s12026-008-8055-8 2009.
  42. Yan, X., Zhao, X., Huo, R., & Xu, T. IRF3 and IRF8 regulate NF-κB signaling by targeting MyD88 in teleost fish. Frontiers in immunology, 11, 606. 2020.
  43. Zhang, R., Chen, K., Peng, L., & Xiong, H. Regulation of T helper cell differentiation by interferon regulatory factor family members. Immunologic research, 54(1), 169-176. https://doi.org/10.1007/s12026-012-8328-0. 2012.
  44. Zhu, K.-C., Guo, H.-Y., Zhang, N., Liu, B.-S., Guo, L., Jiang, S.-G., & Zhang, D.-C. Functional characterization of IRF8 regulation of type II IFN in golden pompano (Trachinotus ovatus). Fish & Shellfish Immunology, 94, 1-9. 2019. https://doi.org/10.1016/j.fsi.2019.08.060