|
|
|
|
|
| Generation and application of two monoclonal antibodies targeting conserved linear epitopes in the NP protein of influenza A virus |
| ZHAO Yu-hui*, WEN Xia*, LI Qi-bing, JIANG Li, WANG Guang-wen, LIANG Li-bin, WANG Xiu-rong, CHEN Hua-lan, LI Cheng-jun |
| State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150069, P.R.China |
|
|
|
Abstract Monoclonal antibodies (mAbs) are widely used in virus research and disease diagnosis. The nucleoprotein (NP) of influenza A virus (IAV) plays important roles in multiple stages of the virus life cycle. Therefore, generating conserved mAbs against NP and characterizing their properties will provide useful tools for IAV research. In this study, two mAbs against the NP protein, 10E9 and 3F3, were generated with recombinant truncated NP proteins (NP-1 and NP-2) as immunogens. The heavy-chain subclass of both 10E9 and 3F3 was determined to be IgG2α, and the light-chain type was κ. Truncation and site-specific mutation analyses showed that the epitopes of mAbs 10E9 and 3F3 were located in the N terminal 84–89 amino acids and the C terminal 320–324 amino acids of the NP protein, respectively. We found that mAbs 10E9 and 3F3 reacted well with the NP protein of H1–H15 subtypes of IAV. Both 10E9 and 3F3 can be used in immunoprecipitation assay, and 10E9 was also successfully applied in confocal microscopy. Furthermore, we found that the 10E9-recognized 84SAGKDP89 epitope and 3F3-recognized 320ENPAH324 epitope were highly conserved in NP among all avian and human IAVs. Thus, the two mAbs we developed could be used as powerful tools in the development of diagnostic methods of IAV, and also surely promote the basic research in understanding the replication mechanisms of IAV.
|
|
Received: 06 October 2021
Accepted: 14 October 2021
|
| Fund: This work was supported by the Natural Science Foundation of Heilongjiang Province, China (JQ2019C005) and the National Natural Science Foundation of China (31702265 and 32172847). |
| About author: Correspondence LI Cheng-jun, Tel: +86-451-51051688, E-mail: lichengjun@caas.cn; CHEN Hua-lan, Tel: +86-451-51997168, E-mail: chenhualan@caas.cn
* These authors contributed equally to this study. |
Cite this article:
ZHAO Yu-hui, WEN Xia, LI Qi-bing, JIANG Li, WANG Guang-wen, LIANG Li-bin, WANG Xiu-rong, CHEN Hua-lan, LI Cheng-jun .
2022.
Generation and application of two monoclonal antibodies targeting conserved linear epitopes in the NP protein of influenza A virus. Journal of Integrative Agriculture, 21(7): 2095-2105.
|
Akarsu H, Burmeister W P, Petosa C, Petit I, Muller C W, Ruigrok R W, Baudin F. 2003. Crystal structure of the M1 protein-binding domain of the influenza A virus nuclear export protein (NEP/NS2). EMBO Journal, 22, 4646–4655.
Arranz R, Coloma R, Chichon F J, Conesa J J, Carrascosa J L, Valpuesta J M, Ortin J, Martin-Benito J. 2012. The structure of native influenza virion ribonucleoproteins. Science, 338, 1634–1637.
Augustin T, Cehlar O, Skrabana R, Majerova P, Hanes J. 2015. Unravelling viral camouflage: Approaches to the study and characterization of conformational epitopes. Acta Virologica, 59, 103–116.
Baudin F, Petit I, Weissenhorn W, Ruigrok R W. 2001. In vitro dissection of the membrane and RNP binding activities of influenza virus M1 protein. Virology, 281, 102–108.
Benjamin D C, Perdue S S. 1996. Site-directed mutagenesis in epitope mapping. Methods, 9, 508–515.
Biswas S K, Boutz P L, Nayak D P. 1998. Influenza virus nucleoprotein interacts with influenza virus polymerase proteins. Journal of Virology, 72, 5493–5501.
Breiteneder H. 2018. Mapping of conformational IgE epitopes of food allergens. Allergy, 73, 2107–2109.
Chan W H, Ng A K, Robb N C, Lam M K, Chan P K, Au S W, Wang J H, Fodor E, Shaw P C. 2010. Functional analysis of the influenza virus H5N1 nucleoprotein tail loop reveals amino acids that are crucial for oligomerization and ribonucleoprotein activities. Journal of Virology, 84, 7337–7345.
Chen H, Yuan H, Gao R, Zhang J, Wang D, Xiong Y, Fan G, Yang F, Li X, Zhou J, Zou S, Yang L, Chen T, Dong L, Bo H, Zhao X, Zhang Y, Lan Y, Bai T, Dong J, et al. 2014. Clinical and epidemiological characteristics of a fatal case of avian influenza A H10N8 virus infection: A descriptive study. Lancet, 383, 714–721.
Chen Y, Zhang J, Qiao C, Wang J, Yang H, Chen H. 2014. Identification of a linear epitope on the haemagglutinin protein of pandemic A/H1N1 2009 influenza virus using monoclonal antibodies. Archives of Virology, 159, 1413–1419.
Gao Y, Huang X, Zhu Y, Lv Z. 2018. A brief review of monoclonal antibody technology and its representative applications in immunoassays. Journal of Immunoassay and Immunochemistry, 39, 351–364.
Garten R J, Davis C T, Russell C A, Shu B, Lindstrom S, Balish A, Sessions W M, Xu X, Skepner E, Deyde V, Okomo-Adhiambo M, Gubareva L, Barnes J, Smith C B, Emery S L, Hillman M J, Rivailler P, Smagala J, de Graaf M, Burke D F, et al. 2009. Antigenic and genetic characteristics of swine-origin 2009 A(H1N1) influenza viruses circulating in humans. Science, 325, 197–201.
Hafeez U, Gan H K, Scott A M. 2018. Monoclonal antibodies as immunomodulatory therapy against cancer and autoimmune diseases. Current Opinion in Pharmacology, 41, 114–121.
He J L, Hsieh M S, Juang R H, Wang C H. 2014. A monoclonal antibody recognizes a highly conserved neutralizing epitope on hemagglutinin of H6N1 avian influenza virus. Veterinary Microbiology, 174, 333–341.
Herz C, Stavnezer E, Krug R, Gurney Jr T. 1981. Influenza virus, an RNA virus, synthesizes its messenger RNA in the nucleus of infected cells. Cell, 26, 391–400.
Hutchinson E C. 2018. Influenza virus. Trends in Microbiology, 26, 809–810.
Hutchinson E C, Denham E M, Thomas B, Trudgian D C, Hester S S, Ridlova G, York A, Turrell L, Fodor E. 2012. Mapping the phosphoproteome of influenza A and B viruses by mass spectrometry. PLoS Pathogens, 8, e1002993.
Jiang L, Fantoni G, Couzens L, Gao J, Plant E, Ye Z, Eichelberger M C, Wan H. 2016. Comparative efficacy of monoclonal antibodies that bind to different epitopes of the 2009 pandemic H1N1 influenza virus neuraminidase. Journal of Virology, 90, 117–128.
Katz C, Levy-Beladev L, Rotem-Bamberger S, Rito T, Rudiger S G, Friedler A. 2011. Studying protein-protein interactions using peptide arrays. Chemical Society Reviews, 40, 2131–2145.
Kaur H, Salunke D M. 2015. Antibody promiscuity: Understanding the paradigm shift in antigen recognition. IUBMB Life, 67, 498–505.
Koopmans M, Wilbrink B, Conyn M, Natrop G, van der Nat H, Vennema H, Meijer A, van Steenbergen J, Fouchier R, Osterhaus A, Bosman A. 2004. Transmission of H7N7 avian influenza A virus to human beings during a large outbreak in commercial poultry farms in the Netherlands. Lancet, 363, 587–593.
Li C, Chen H. 2014. Enhancement of influenza virus transmission by gene reassortment. Current Topics in Microbiology and Immunology, 385, 185–204.
Li C J, Chen H L. 2021. H7N9 influenza virus in China. Cold Spring Harbor Perspectives in Medicine, 11, a038349.
Liang L B, Jiang L, Li J P, Zhao Q Q, Wang J G, He X J, Huang S Y, Wang Q, Zhao Y H, Wang G W, Sun N, Deng G H, Shi J Z, Tian G B, Zeng X Y, Jiang Y P, Liu L L, Liu J X, Chen P C, Bu Z G, et al. 2019. Low Polymerase Activity Attributed to PA Drives the Acquisition of the PB2 E627K Mutation of H7N9 Avian Influenza Virus in Mammals. mBio, 10, e01162–e01181.
Liu Z, Chen Z, Hong J, Wang X, Zhou C, Zhou X, Wu J. 2016. Monoclonal antibody-based serological methods for detecting Citrus tristeza virus in citrus groves. Virologica Sinica, 31, 324–330.
Luo W, Zhang J, Liang L, Wang G, Li Q, Zhu P, Zhou Y, Li J, Zhao Y, Sun N, Huang S, Zhou C, Chang Y, Cui P, Chen P, Jiang Y, Deng G, Bu Z, Li C, Jiang L, et al. 2018. Phospholipid scramblase 1 interacts with influenza A virus NP, impairing its nuclear import and thereby suppressing virus replication. PLoS Pathogens, 14, e1006851.
Marklund J K, Ye Q Z, Dong J H, Tao Y J, Krug R M. 2012. Sequence in the influenza A virus nucleoprotein required for viral polymerase binding and RNA synthesis. Journal of Virology, 86, 7292–7297.
Moeller A, Kirchdoerfer R N, Potter C S, Carragher B, Wilson I A. 2012. Organization of the influenza virus replication machinery. Science, 338, 1631–1634.
Moreira G, Fuhner V, Hust M. 2018. Epitope mapping by phage display. Methods in Molecular Biology, 1701, 497–518.
Neumann G, Chen H, Gao G F, Shu Y L, Kawaoka Y. 2010. H5N1 influenza viruses: Outbreaks and biological properties. Cell Research, 20, 51–61.
Neumann G, Noda T, Kawaoka Y. 2009. Emergence and pandemic potential of swine-origin H1N1 influenza virus. Nature, 459, 931–939.
Neumann G, Watanabe T, Ito H, Watanabe S, Goto H, Gao P, Hughes M, Perez D R, Donis R, Hoffmann E, Hobom G, Kawaoka Y. 1999. Generation of influenza A viruses entirely from cloned cDNAs. Proceedings of the National Academy of Sciences of the United States of America, 96, 9345–9350.
Noton S L, Medcalf E, Fisher D, Mullin A E, Elton D, Digard P. 2007. Identification of the domains of the influenza A virus M1 matrix protein required for NP binding, oligomerization and incorporation into virions. Journal of General Virology, 88, 2280–2290.
Paterson D, Fodor E. 2012. Emerging roles for the influenza A virus nuclear export protein (NEP). PLoS Pathogens, 8, e1003019.
Pleschka S, Jaskunas S R, Engelhardt O G, Zurcher T, Palese P, GarciaSastre A. 1996. A plasmid-based reverse genetics system for influenza A virus. Journal of Virology, 70, 4188–4192.
Shao W, Li X, Goraya M U, Wang S, Chen J L. 2017. Evolution of influenza A virus by mutation and re-assortment. International Journal of Molecular Sciences, 18, 1650.
Shi J Z, Deng G H, Kong H H, Gu C Y, Ma S J, Yin X, Zeng X Y, Cui P F, Chen Y, Yang H L, Wan X P, Wang X R, Liu L L, Chen P C, Jiang Y P, Liu J X, Guan Y T, Suzuki Y, Li M, Qu Z Y, et al. 2017. H7N9 virulent mutants detected in chickens in China pose an increased threat to humans. Cell Research, 27, 1409–1421.
Shi J Z, Deng G H, Ma S J, Zeng X Y, Yin X, Li M, Zhang B, Cui P F, Chen Y, Yang H L, Wan X P, Liu L L, Chen P C, Jiang Y P, Guan Y T, Liu J X, Gu W L, Han S Y, Song Y M, Liang L B, et al. 2018. Rapid evolution of H7N9 highly pathogenic viruses that emerged in China in 2017. Cell Host & Microbe, 24, 558–568.
Shimizu T, Takizawa N, Watanabe K, Nagata K, Kobayashi N. 2011. Crucial role of the influenza virus NS2 (NEP) C-terminal domain in M1 binding and nuclear export of vRNP. FEBS Letters, 585, 41–46.
Singh P, Toom S, Huang Y. 2017. Anti-claudin 18.2 antibody as new targeted therapy for advanced gastric cancer. Journal of Hematology & Oncology, 10, 105.
Steentoft C, Fuhrmann M, Battisti F, Van Coillie J, Madsen T D, Campos D, Halim A, Vakhrushev S Y, Joshi H J, Schreiber H, Mandel U, Narimatsu Y. 2019. A strategy for generating cancer-specific monoclonal antibodies to aberrant O-glycoproteins: Identification of a novel dysadherin-Tn antibody. Glycobiology, 29, 307–319.
Subbarao K. 2019. The critical interspecies transmission barrier at the animal-human interface. Tropical Medicine and Infectious Disease, 4, 72.
Sun Y P, Liu J H. 2015. H9N2 influenza virus in China: A cause of concern. Protein & Cell, 6, 18–25.
Sun Z, Shi B, Meng F, Ma R, Hu Q, Qin T, Chen S, Peng D, Liu X. 2018. Development of a colloidal gold-based immunochromatographic strip for rapid detection of H7N9 influenza viruses. Frontiers in Microbiology, 9, 2069.
Tong S, Zhu X, Li Y, Shi M, Zhang J, Bourgeois M, Yang H, Chen X, Recuenco S, Gomez J, Chen L M, Johnson A, Tao Y, Dreyfus C, Yu W, McBride R, Carney P J, Gilbert A T, Chang J, Guo Z, et al. 2013. New world bats harbor diverse influenza A viruses. PLoS Pathogens, 9, e1003657.
Tong X C, Weng S S, Xue F, Wu X, Xu T M, Zhang W H. 2018. First human infection by a novel avian influenza A(H7N4) virus. Journal of Infection, 77, 255–257.
Tu Y C, Chen K Y, Chen C K, Cheng M C, Lee S H, Cheng I C. 2019. Novel application of influenza A virus-inoculated chorioallantoic membrane to characterize a NP-specific monoclonal antibody for immunohistochemistry assaying. Journal of Veterinary Science, 20, 51.
Tweed S A, Skowronski D M, David S T, Larder A, Petric M, Lees W, Li Y, Katz J, Krajden M, Tellier R, Halpert C, Hirst M, Astell C, Lawrence D, Mak A. 2004. Human illness from avian influenza H7N3, British Columbia. Emerging Infectious Diseases, 10, 2196–2199.
Wan H, Yang H, Shore D A, Garten R J, Couzens L, Gao J, Jiang L, Carney P J, Villanueva J, Stevens J, Eichelberger M C. 2015. Structural characterization of a protective epitope spanning A(H1N1)pdm09 influenza virus neuraminidase monomers. Nature Communications, 6, 6114.
Webster R G, Bean W J, Gorman O T, Chambers T M, Kawaoka Y. 1992. Evolution and ecology of influenza A viruses. Microbiological Reviews, 56, 152–179.
Yamanaka K, Ishihama A, Nagata K. 1990. Reconstitution of influenza virus RNA-nucleoprotein complexes structurally resembling native viral ribonucleoprotein cores. Journal of Biological Chemistry, 265, 11151–11155.
Yang Z F, Mok C K, Peiris J S, Zhong N S. 2015. Human infection with a novel avian influenza A(H5N6) virus. New England Journal of Medicine, 373, 487–489.
Ye Q Z, Krug R M, Tao Y Z J. 2006. The mechanism by which influenza A virus nucleoprotein forms oligomers and binds RNA. Nature, 444, 1078–1082.
|
| No Suggested Reading articles found! |
|
|
Viewed |
|
|
|
Full text
|
|
|
|
|
Abstract
|
|
|
|
|
Cited |
|
|
|
|
| |
Shared |
|
|
|
|
| |
Discussed |
|
|
|
|
