Multiple microRNAs targeted to internal ribosome entry site against foot-and-mouth disease virus infection in vitro and in vivo

Virology Journal

Xuepeng Cai²  Xiangtao Liu³  Zaixin Liu³  Kebin Mu¹  Xuerong Liu¹  Xuenong Luo³  Huifang Bao³  Yongxi Dou³  Yanyan Chang¹ 

[1] Research & Development Department, China Agricultural Veterinary Biological Science and Technology Co., LTD, Lanzhou 730046, Gansu, P. R. of China;China Animal Disease Prevention Control Center, Beijing 100125, P. R. of China;State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, Gansu, P. R of China

关键词: Real-time quantitative RT-PCR;    Flow cytometry;    Antiviral effect;    Transformed cell clones;    Internal ribosome entry site;    MicroRNA;    Foot-and-mouth disease virus;   

Others  :  819914
DOI  :  10.1186/1743-422X-11-1

 received in 2013-09-22, accepted in 2013-12-27,  发布年份 2014

【 摘 要 】

Background

Foot-and-mouth disease virus (FMDV) causes a severe vesicular disease in domestic and wild cloven-hoofed animals. Because of the limited early protection induced by current vaccines, emergency antiviral strategies to control the rapid spread of FMD outbreaks are needed.

Here we constructed multiple microRNAs (miRNAs) targeting the internal ribosome entry site (IRES) element of FMDV and investigated the effect of IRES-specific miRNAs on FMDV replication in baby hamster kidney (BHK-21) cells and suckling mice.

Results

Four IRES-specific miRNAs significantly reduced enhanced green fluorescent protein (EGFP) expression from IRES-EGFP reporter plasmids, which were used with each miRNA expression plasmid in co-transfection of BHK-21 cells. Furthermore, treatment of BHK-21 cells with Bi-miRNA (a mixture of two miRNA expression plasmids) and Dual-miRNA (a co-cistronic expression plasmid containing two miRNA hairpin structures) induced more efficient and greater inhibition of EGFP expression than did plasmids carrying single miRNA sequences.

Stably transformed BHK-21 cells and goat fibroblasts with an integrating IRES-specific Dual-miRNA were generated, and real-time quantitative RT-PCR showed that the Dual-miRNA was able to effectively inhibit the replication of FMDV (except for the Mya98 strain) in the stably transformed BHK-21 cells.

The Dual-miRNA plasmid significantly delayed the deaths of suckling mice challenged with 50× and 100× the 50% lethal dose (LD₅₀) of FMDV vaccine strains of three serotypes (O, A and Asia 1), and induced partial/complete protection against the prevalent PanAsia-1 and Mya98 strains of FMDV serotype O.

Conclusion

These data demonstrate that IRES-specific miRNAs can significantly inhibit FMDV infection in vitro and in vivo.

【 授权许可】

   
2013 Chang et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Grubman MJ, Baxt B: Foot-and-mouth disease. Clin Microbiol Rev 2004, 17:465-493.
• [2]Alexandersen S, Mowat N: Foot-and-mouth disease: host range and pathogenesis. Curr Top Microbiol Immunol 2005, 288:9-42.
• [3]Belsham GJ: Distinctive features of foot-and-mouth disease virus, a member of the picornavirus family: aspects of virus protein synthesis, protein processing and structure. Prog Biolphys Mol Biol 1993, 69:241-260.
• [4]Bachrach HL: Foot-and-mouth disease. Annu Rev Microbiol 1968, 22:201-244.
• [5]Brown F: The history of research in foot-and-mouth disease. Virus Res 2003, 91:3-7.
• [6]Kitching RP: Global epidemiology and prospects for control of foot-and-mouth disease. Curr Microbiol Immunol 2005, 288:133-148.
• [7]Sobrino F, Saiz M, Jimenez-clavero MA, Nunez JI, Rosas MF, Baranowski E, Ley V: Foot-and-mouth disease virus: a long known virus, but a current threat. Vet Res 2001, 32:1-30.
• [8]Mason PW, Grubman MJ, Baxt B: Molecular basis of pathogenesis of FMDV. Virus Res 2003, 91:9-32.
• [9]Ryan MD, Donelly MLL, Flint M, Cowton VM, Luke G, Hughes LE, Knox C, De Felipe P: Foot-and-mouth disease virus proteinases. In Foot-and Mouth Disease Current Perspectives. Edited by Sobrino F, Domingo E. London: Horizon Scientific Press; 2004:53-76.
• [10]Belsham GJ, Brangwyn JK: A region of the 5′ non-coding region of foot-and-mouth disease virus RNA directs efficient internal initiation of protein synthesis within cells; interaction with the role of the L protease in translational control. J Virol 1990, 64:5389-5395.
• [11]Kuhn R, Luz N, Beck E: Functional analysis of the internal translation initiation site of foot-and-mouth disease virus. J Virol 1990, 64:4625-4631.
• [12]Pilipenko EV, Blinov VM, Chernov BK, Dmitrieva TM, Agol VI: Conservation of the secondary structure elements of the 5′-untranslated region of cardiovirus and aphthovirus RNAs. Nucl Acids Res 1989, 17:5701-5711.
• [13]Belsham GJ, Martinez-Salas E: Genome organization, translation and replication of foot-and-mouth disease virus RNA. In Foot-and-Mouth Disease Current Perspectives. Edited by Sobrino F, Domingo E. London: Horizon Scientific Press; 2004:19-52.
• [14]Belsham GJ: Translation and replication of FMDV RNA. Curr Top Microbiol Immunol 2005, 288:43-70.
• [15]Domingo E, Ruiz-Jarabo CM, Sierra S, Arias A, Pariente N, Baranowski E, Escarmis C: Emergence and selection of RNA virus variant: memory and extinction. Virus Res 2001, 82:39-44.
• [16]Domingo E, Baranowski E, Escarmis C, Sobrino F, Holland JJ: Error frequencies of picornavirus RNA polymerases: evolutionary implications for viral populations. In Molecular Biology of Picornaviruses. Edited by Semler BL, Wimmer E. Washington: DC: ASM Press; 2002:285-298.
• [17]Cottam EM, Haydon DT, Paton DJ, Gloster J, Wilesmith JW, Ferris NP, Hutchings GH, King DP: Molecular epidemiology of the foot-and-mouth disease virus outbreak in the United Kingdom in 2001. J Virol 2006, 80:11274-11282.
• [18]Martinez MA, Carrillo C, Gonzalez-Candelas F, Moya A, Domingo E, Sobrino F: Fitness alteration of foot-and-mouth disease virus mutants: measurement of adaptability of viral quasispecies. J Virol 1991, 65:3954-3957.
• [19]Domingo E, Escarmis C, Martinez MA, Martinez-Salas E, Mateu MG: Foot-and-mouth disease virus populations are quasispecies. Curr Top Microbiol Immunol 1992, 176:33-47.
• [20]Ruiz-Jarabo CM, Arias M, Baranowski E, Escarmis C, Domingo E: Memory in viral quasispecies. J Virol 2000, 74:3543-3547.
• [21]Domingo E, Pariente N, Airaksinen A, Gonzalez-Lopez C, Sierra S, Herrera M, Grande-Perez A, Lowenstein PR, Manrubia SC, Lazaro E, Escarmis C: Foot-and-mouth disease virus evolution: exploring pathways towards virus extinction. populations are quasispecies. Curr Top Microbiol Immunol 2005, 288:149-173.
• [22]Domingo E, Escarmis C, Baranowski E, Ruiz-Jarabo CM, Carrillo E, Nunez JI, Sobrino F: Evolution of foot-and-mouth disease virus. Virus Res 2003, 91:47-63.
• [23]Domingo E, Escarmis C, Lazaro E, Manrubia SC: Quasispecies dynamics and RNA virus extinction. Virus Res 2005, 107:129-139.
• [24]Doel TR: FMD vaccines. Virus Res 2003, 91:81-99.
• [25]Yang B, Lan X, Li X, Yin X, Li B, Han X, Li Y, Zhang Z, Liu J: A novel bi-functional DNA vaccine expressing VP1 protein and producing antisense RNA targeted to 5′UTR of foot-and-mouth disease virus can induce both rapid inhibitory effect and specific immune response in mice. Vaccine 2008, 26:5477-5483.
• [26]Chen W, Yan W, Du Q, Fei L, Liu M, Ni Z, Sheng Z, Zheng Z: RNA interference targeting VP1 inhibits foot-and-mouth disease virus replication in BHK-21 cells and suckling mice. J Virol 2004, 78:6900-6907.
• [27]Kahana R, Kuznetzova L, Rogel A, Shemesh M, Hai D, Yadin H, Stram Y: Inhibition of foot-and-mouth disease virus replication by small interfering RNA. J Gen Virol 2004, 85:3213-3217.
• [28]Grubman MJ, De los Santos T: Rapid control of foot-and-mouth disease outbreaks: is RNAi a possible solution? Trends Immunol 2005, 26:65-68.
• [29]Chen W, Liu M, Jiao Y, Yan W, Wei X, Chen J, Fei L, Liu Y, Zuo X, Yang F, Lu Y, Zheng Z: Adenovirus-mediated RNA interference against foot-and-mouth disease virus infection both in vitro and in vivo. J Virol 2006, 80:3559-3566.
• [30]Kim SM, Lee KN, Park JY, Ko YJ, Joo YS, Kim HS, Park JH: Therapeutic application of RNA interference against foot-and-mouth disease virus in vitro and in vivo. Antiviral Res 2008, 80:178-184.
• [31]Wang PY, Ren Y, Guo ZR, Chen CF: Inhibition of foot-and-mouth disease virus replication in vitro and in vivo by small interfering RNA. Virol J 2008, 5:86. BioMed Central Full Text
• [32]Lv K, Guo YJ, Zhang YL, Wang KY, Li K, Zhu Y, Sun SH: Transient inhibition of foot-and-mouth disease virus replication by siRNAs silencing VP1 protein coding region. Res Vet Sci 2009, 86:443-452.
• [33]Cong W, Cui SQ, Chen JL, Zuo XP, Lu YG, Yan WY, Zheng ZX: Construction of a multiple targeting RNAi plasmid that inhibits target gene expression and FMDV replication in BHK-21 cells and suckling mice. Vet Res Commun 2010, 34:335-346.
• [34]Boden D, Pusch O, Lee F, Tucker L, Ramratnam B: Human immunodeficiency virus type 1 escape from RNA interference. J Virol 2003, 77:11531-11535.
• [35]Pusch O, Boden D, Silbermann R, Lee F, Tucker L, Ramratnam B: Nucleotide sequence homology requirements of HIV-1-specific short hairpin RNA. Nucleic Acids Res 2003, 31:6444-6449.
• [36]Gitlin L, Stone JK, Andino R: Poliovirus escape from RNA interference: short interfering RNA-target recognition and implications for therapeutic approaches. J Virol 2005, 79:1027-1035.
• [37]Sabariegos R, Giménez-Barcons M, Tàpia N, Clotet B, Martínez MA: Sequence homology required by human immunodeficiency virus type 1 to escape from short interfering RNAs. J Virol 2006, 80:571-577.
• [38]Kim SM, Lee KN, Lee SJ, Ko YJ, Lee HS, Kweon CH, Kim HS, Park JH: Multiple shRNAs driven by U6 and CMV promoter enhances efficiency of antiviral effects against foot-and-mouth disease virus. Antiviral Res 2010, 87:307-317.
• [39]Nilsen TW: Mechanisms of microRNA-mediated gene regulation in animal cells. Trends Genet 2007, 23:243-249.
• [40]Filipowicz W, Bhattacharyya SN, Sonenberg N: Mechanisms of post-transcriptional regulation by microRNAs: are the answers in sight? Nat Rev Genet 2008, 9:102-114.
• [41]Du JZ, Gao SD, Luo JH, Zhang GF, Cong GZ, Shao JJ, Lin T, Cai XP, Chang HY: Effective inhibition of foot-and-mouth disease virus (FMDV) replication in vitro by vector-delivered microRNAs targeting the 3D gene. Virol J 2011, 8:292. BioMed Central Full Text
• [42]Carrington JC, Ambros V: Role of microRNAs in plant and animal development. Science 2003, 301:336-338.
• [43]Pillai RS, Bhattacharyya SN, Filipowicz W: Repression of protein synthesis by miRNAs: how many mechanisms? Trends Cell Biol 2007, 17:118-126.
• [44]Bartel DP: MicroRNAs: target recognition and regulatory functions. Cell 2009, 136:215-233.
• [45]He L, Hannon GJ: MicroRNAs: small RNAs with a big role in gene regulation. Nat Rev Genet 2004, 5:522-531.
• [46]Sonkoly E, Pivarcsi A: Advances in microRNAs: implications for immunity and inflammatory diseases. J Cell Mol Med 2009, 13:24-38.
• [47]Fernández-Miragall O, Martínez-Salas E: Structural organization of a viral IRES depends on the integrity of the GNRA motif. RNA 2003, 9:1333-1344.
• [48]Drew J, Belsham GJ: Trans-complementation by RNA of defective FMDV internal ribosome entry site elements. J Virol 1994, 68:697-703.
• [49]Ramos R, Martinez-Salas E: Long-range RNA interactions between strutural domains of the aphthovirus internal ribosome entry site (IRES). RNA 1999, 5:1374-1383.
• [50]Roberts LO, Belsham GJ: Complementation of defective picornavirus internal ribosome entry site (IRES) elements by the coexpression of fragments of the IRES. Virology 1997, 227:53-62.
• [51]Gutiérrez A, Martínez-Salas E, Pintado B, Sobrino F: Specific inhibition of aphthovirus infection by RNAs transcribed from both the 5′ and the 3′ noncoding regions. J Virol 1994, 68:7426-7432.
• [52]Bigeriego P, Rosas MF, Zamora E, Martínez-Salas E, Sobrino F: Heterotypic inhibition of foot-and-mouth disease virus infection by combinations of RNA transcripts corresponding to the 5′ and 3′ regions. Antiviral Res 1999, 44:133-141.
• [53]Rosas MF, Martínez-Salas E, Sobrino F: Stable expression of antisense RNAs targeted to the 5′ non-coding region confers heterotypic inhibition to foot-and-mouth disease virus infection. J Gen Virol 2003, 84:393-402.
• [54]Rodríguez-Pulido M, Sobrino F, Borrego B, Sáiz M: Inoculation of newborn mice with non-coding regions of foot-and-mouth disease virus RNA can induce a rapid, solid and wide-range protection against viral infection. Antiviral Res 2011, 92:500-504.
• [55]Joyappa DH, Sasi S, Ashok KC, Reddy GR, Suryanarayana VV: The plasmid constructs producing shRNA corresponding to the conserved 3D polymerase of foot and mouth disease virus protects guinea pigs against challenge virus. Vet Res Commun 2009, 33:263-271.
• [56]De Los Santos T, Wu Q, De Avila Botton S, Grubman MJ: Short hairpin RNA targeted to the highly conserved 2B nonstructural protein coding region inhibits replication of multiple serotypes of foot-and-mouth disease virus. Virology 2005, 335:222-231.
• [57]Stassinopoulos IA, Belsham GJ: A novel protein-RNA binding assay: functional interactions of the foot-and-mouth disease virus internal ribosome entry site with cellular proteins. RNA 2001, 7:114-122.
• [58]Martínez-Salas E: The impact of RNA structure on picornavirus IRES activity. Trends Microbiol 2008, 16:230-237.
• [59]Wilson JA, Richardson CD: Hepatitis C virus replicons escape RNA interference induced by a short interfering RNA directed against the NS5b coding region. J Virol 2005, 79:7050-7058.
• [60]Bai X, Li P, Bao H, Liu Z, Li D, Lu Z, Cao Y, Shang Y, Shao J, Chang H, Luo J, Liu X: Evolution and molecular epidemiology of foot-and-mouth disease virus in China. Chin Sci Bull 2011, 56:2191-2201.
• [61]Cong W, Jin H, Jiang C, Yan W, Liu M, Chen J, Zuo X, Zheng Z: Attenuated Salmonella choleraesuis-mediated RNAi targeted to conserved regions against foot-and-mouth disease virus in guinea pigs and Swine. Vet Res 2010, 41:30.
• [62]Bao HF, Li D, Sun P, Zhou Q, Hu J, Bai XW, Fu YF, Lu ZJ, Liu ZX: The infectivity and pathogenicity of a foot-and-mouth disease virus persistent infection strain from oesophageal-pharyngeal fluid of a Chinese cattle in 2010. Virol J 2011, 8:536. BioMed Central Full Text
• [63]Escarmís C, Carrillo EC, Ferrer M, Arriaza JF, Lopez N, Tami C, Verdaguer N, Domingo E, Franze-Fernández MT: Rapid selection in modified BHK-21 cells of a foot-and-mouth disease virus variant showing alterations in cell tropism. J Virol 1998, 72:10171-10179.
• [64]Vagnozzi A, Stein DA, Iversen PL, Rieder E: Inhibition of foot-and-mouth disease virus infections in cell cultures with antisense morpholino oligomers. J Virol 2007, 81:11669-11680.
• [65]Fajardo T Jr, Rosas MF, Sobrino F, Martinez-Salas E: Exploring IRES region accessibility by interference of foot-and-mouth disease virus infectivity. PLoS One 2012, 7:e41382.
• [66]Norder H, De Palma AM, Selisko B, Costenaro L, Papageorgiou N, Arnan C, Coutard B, Lantez V, De Lamballerie X, Baronti C, Solà M, Tan J, Neyts J, Canard B, Coll M, Gorbalenya AE, Hilgenfeld R: Picornavirus non-structural proteins as targets for new anti-virals with broad activity. Antiviral Res 2011, 89:204-218.
• [67]Cao YM, Lu ZJ, Sun P, Fu YF, Tian FP, Hao XF, Bao HF, Liu XT, Liu ZX: A pseudotype baculovirus expressing the capsid protein of foot-and-mouth disease virus and a T-cell immunogen shows enhanced immunogenicity in mice. Virol J 2011, 8:77. BioMed Central Full Text
• [68]Liu XS, Wang YL, Zhang YG, Fang YZ, Pan L, Lu JL, Zhou P, Zhang ZW, Jiang ST: Identification of H-2d restricted T cell epitope of foot-and-mouth disease virus structural protein VP1. Virol J 2011, 8:426. BioMed Central Full Text