禽流感病毒RNA聚合酶真核表达纯化及与其结合宿主因子的初步研究
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摘要
在本研究中,我们利用原核表达的流感病毒RNA聚合酶碱性蛋白2(polymerase basic protein 2,PB2)免疫BABL/c小鼠,经过细胞融合与杂交瘤细胞筛选制备针对流感病毒聚合酶亚基PB2的单克隆抗体5G11。实验表明,5G11可以与H5亚型不同毒株的PB2蛋白反应。经单抗亚型鉴定,5G11的重链为IgG2a,轻链为κ链。利用这株单抗从病毒感染细胞的裂解上清中,免疫共沉淀得到了病毒的RNA聚合酶复合体。通过二维电泳区分免疫共沉淀得到的蛋白混合物,找到并经质谱鉴定了可能与病毒RNA聚合酶相结合的宿主蛋白;此外,我们构建了带有TAP标签的流感病毒聚合酶三个亚基的真核表达质粒:pCDNA-35PB1、pCDNA-35PB2TAP和pCDNA-35PA。将这三种质粒转染293T细胞后,经过串联亲和纯化方法得到流感病毒聚合酶复合体。
本试验成功建立了流感病毒RNA聚合酶的真核表达及纯化方法,得到了真核细胞内表达的流感病毒RNA聚合酶复合体,初步研究了与流感病毒聚合酶相作用的宿主相关蛋白,为研究二者间相互作用在病毒致病机理中作用奠定了基础,有利于我们进一步了解流感病毒致病机理,也有利于新型抗流感病毒药物的研制。
In this study, the RNA-dependent RNA polymerase (RdRp) of avian influenza virus (AIV) was expressed and purified from eukaryotic cells in two ways.One is developing one hybridoma cell strain named 5G11 by confusing SP2/0 cells and spleen cells of mouse immunized with prokaryotic expressed PB2 protein . The results of Western-blot indicated that 5G11 can react with seavral H5 subtype strains’PB2 protein. The results of subtype analysis showed that the heavy chain of 5G11 belonged to IgG2a and the light chain was theκchain. two different H5 subtype AIV strains’RdRp can be purified from virus infected MDCK with 5G11 by co-immunoprecipitation assay,although these two stains possess different virulence to mouse.we found and identified several proteins that may sociate with virus RdRp by 2-D electrophoresis and mass spectrographic analysis. The other method is that we constructed eurokaryotic expression reconstructed plasmids of RdRp,with TAP-tag in the fused protein PB2 subunit.Then,by tandem affinity purification procedure (TAP), virus RdRp was pufied from plasmids cotransfected 293T cells.
In summary, we have set up reliable method of expression and purification of AIV’s RdRp in eurokaryotic cells and we purified virus RdRp from eurokaryotic cell.Based on this, the host factors combining with virus RdRp was researched preliminarily.All these work together provides an efficient avenue to study the host factors’function in AIV replicating in mammalian cells.This work helps us to understand better the pathogenicity machnism of influenza virus and to go a step further ,to develop new drugs with high efficiency in future.
本试验成功建立了流感病毒RNA聚合酶的真核表达及纯化方法,得到了真核细胞内表达的流感病毒RNA聚合酶复合体,初步研究了与流感病毒聚合酶相作用的宿主相关蛋白,为研究二者间相互作用在病毒致病机理中作用奠定了基础,有利于我们进一步了解流感病毒致病机理,也有利于新型抗流感病毒药物的研制。
In this study, the RNA-dependent RNA polymerase (RdRp) of avian influenza virus (AIV) was expressed and purified from eukaryotic cells in two ways.One is developing one hybridoma cell strain named 5G11 by confusing SP2/0 cells and spleen cells of mouse immunized with prokaryotic expressed PB2 protein . The results of Western-blot indicated that 5G11 can react with seavral H5 subtype strains’PB2 protein. The results of subtype analysis showed that the heavy chain of 5G11 belonged to IgG2a and the light chain was theκchain. two different H5 subtype AIV strains’RdRp can be purified from virus infected MDCK with 5G11 by co-immunoprecipitation assay,although these two stains possess different virulence to mouse.we found and identified several proteins that may sociate with virus RdRp by 2-D electrophoresis and mass spectrographic analysis. The other method is that we constructed eurokaryotic expression reconstructed plasmids of RdRp,with TAP-tag in the fused protein PB2 subunit.Then,by tandem affinity purification procedure (TAP), virus RdRp was pufied from plasmids cotransfected 293T cells.
In summary, we have set up reliable method of expression and purification of AIV’s RdRp in eurokaryotic cells and we purified virus RdRp from eurokaryotic cell.Based on this, the host factors combining with virus RdRp was researched preliminarily.All these work together provides an efficient avenue to study the host factors’function in AIV replicating in mammalian cells.This work helps us to understand better the pathogenicity machnism of influenza virus and to go a step further ,to develop new drugs with high efficiency in future.
引文
1. B.W.卡尔尼克.禽病学(高福,苏敬良主译).(第十版)[M].北京:中国农业出版社,1999,742-771.
2.关薇,王建,贺福初.大规模蛋白质相互作用研究方法进展[J].生命科学,2006,18(5),507-511.
3.葛叶,邓国华,李雪平,陈化兰.流感病毒PB2亚单位抗血清的制备[J].中国预防兽医学报,已接受.
4.葛叶,邓国华,李雪平,陈化兰.流感病毒聚合酶PB1-1蛋白的表达与单克隆抗体的制备[J].中国预防兽医学报,2009,(9),726-729.
5.葛叶,邓国华,李雪平,高玉伟,姚秋成,唐艳玲,陈化兰.流感病毒聚合酶PA亚单位抗血清的制备[J].中国预防兽医学报,2009,(4),314-317.
6.童光志,王云峰.动物基因工程疫苗原理与方法[M].北京:化学工业出版社,2009,30-40
7.王海波,安学丽,张艳贞,王爱丽,李巧云,晏月明.蛋白质相互作用研究方法及其应用[J].生物技术通报,2006年增刊,167-170.
8.张成裕.免疫增强剂在动物医学上的研究进展及应用[J].福建畜牧兽医,2004(1),11-12.
9. Alexander, D.J. and Brown, I.H. Recent zoonoses caused by influenza A viruses[J]. Rev Sci Tech ,2000,19(1), 197-225.
10. Area, E., Martin-Benito, J., Gastaminza, P., Torreira, E., Valpuesta, J.M., Carrascosa, J.L. and Ortin, J. 3D structure of the influenza virus polymerase complex: localization of subunit domains[J]. Proc Natl Acad Sci U S A ,2004,101(1), 308-13.
11. Basler, C.F. and Aguilar, P.V. Progress in identifying virulence determinants of the 1918 H1N1 and the Southeast Asian H5N1 influenza A viruses[J]. Antiviral Res ,2008,79(3) 166-78.
12. Biswas, S.K. and Nayak, D.P. Mutational analysis of the conserved motifs of influenza A virus polymerase basic protein 1[J]. J Virol 6,1994,8(3), 1819-26.
13. Boulo, S., Akarsu, H., Ruigrok, R.W. and Baudin, F. Nuclear traffic of influenza virus proteins and ribonucleoprotein complexes[J]. Virus Res ,2007,124(1-2), 12-21.
14. Burgui, I., Yanguez, E., Sonenberg, N. and Nieto, A. Influenza virus mRNA translation revisited: is the eIF4E cap-binding factor required for viral mRNA translation? [J] J Virol ,2007,81(22), 12427-38.
15. Cardona, C.J., Xing, Z., Sandrock, C.E. and Davis, C.E. Avian influenza in birds and mammals[J]. Comp Immunol Microbiol Infect Dis ,2009,32(4), 255-73.
16. Chan, A.Y., Vreede, F.T., Smith, M., Engelhardt, O.G. and Fodor, E. Influenza virus inhibits RNA polymerase II elongation[J]. Virology ,2006,351(1), 210-7.
17. Deng, T., Engelhardt, O.G., Thomas, B., Akoulitchev, A.V., Brownlee, G.G. and Fodor, E. Role of ran binding protein 5 in nuclear import and assembly of the influenza virus RNA polymerase complex[J]. J Virol ,2006,80(24), 11911-9.
18. Desselberger, U., Racaniello, V.R., Zazra, J.J. and Palese, P. , The 3' and 5'-terminal sequences of influenza A, B and C virus RNA segments are highly conserved and show partial inverted complementarity[J]. Gene,1980, 8(3), 315-28.
19. Engelhardt, O.G., Smith, M. and Fodor, E. Association of the influenza A virus RNA-dependent RNA polymerase with cellular RNA polymerase II[J]. J Virol ,2005,79(9), 5812-8.
20. Fechter, P., Mingay, L., Sharps, J., Chambers, A., Fodor, E. and Brownlee, G.G. Two aromatic residues in the PB2 subunit of influenza A RNA polymerase are crucial for cap binding[J]. J Biol Chem ,2003,278(22), 20381-8.
21. Flick, R. and Hobom, G. Interaction of influenza virus polymerase with viral RNA in the 'corkscrew' conformation[J]. J Gen Virol ,1999,80 (Pt 10), 2565-72.
22. Fodor, E., Pritlove, D.C. and Brownlee, G.G. Characterization of the RNA-fork model of virion RNA in the initiation of transcription in influenza A virus[J]. J Virol ,1995,69(7), 4012-9.
23. Fujii, K., Fujii, Y., Noda, T., Muramoto, Y., Watanabe, T., Takada, A., Goto, H., Horimoto, T. and Kawaoka, Y. Importance of both the coding and the segment-specific noncoding regions of the influenza A virus NS segment for its efficient incorporation into virions[J]. J Virol, 2005, 79(6), 3766-74.
24. Gabriel, G., Abram, M., Keiner, B., Wagner, R., Klenk, H.D. and Stech, J. Differential polymerase activity in avian and mammalian cells determines host range of influenza virus[J]. J Virol, 2007, 81(17), 9601-4.
25. Gabriel, G., Herwig, A. and Klenk, H.D. Interaction of polymerase subunit PB2 and NP with importin alpha1 is a determinant of host range of influenza A virus[J]. PLoS Pathog, 2008, 4(2), e11.
26. Hara, K., Schmidt, F.I., Crow, M. and Brownlee, G.G. Amino acid residues in the N-terminal region of the PA subunit of influenza A virus RNA polymerase play a critical role in protein stability, endonuclease activity, cap binding, and virion RNA promoter binding[J]. J Virol, 2006,80(16), 7789-98.
27. Honda, A., Mizumoto, K. and Ishihama, A. Two separate sequences of PB2 subunit constitute the RNA cap-binding site of influenza virus RNA polymerase[J]. Genes Cells ,1999,4(8), 475-85.
28. Hossain, M.J., Mori, I., Dong, L., Liu, B. and Kimura, Y. Fetal calf serum inhibits virus genome expression in Madin-Darby canine kidney cells persistently infected with influenza A virus[J]. Med Microbiol Immunol, 2008, 197(1), 21-7.
29. Hsu, M.T., Parvin, J.D., Gupta, S., Krystal, M. and Palese, P. Genomic RNAs of influenza viruses are held in a circular conformation in virions and in infected cells by a terminal panhandle[J]. Proc Natl Acad Sci U S A, 1987, 84(22), 8140-4.
30. Huarte, M., Sanz-Ezquerro, J.J., Roncal, F., Ortin, J. and Nieto, A. PA subunit from influenza virus polymerase complex interacts with a cellular protein with homology to a family of transcriptional activators[J]. J Virol, 2001, 75(18), 8597-604.
31. Hynes RO. The extracellular matrix: not just pretty fibrils[J].Science, 2009, 326(5957),1216-9.
32. Ito, T. and Kawaoka, Y. Host-range barrier of influenza A viruses[J]. Vet Microbiol, 2000, 74(1-2), 71-5.
33. Jones, I.M., Reay, P.A. and Philpott, K.L. Nuclear location of all three influenza polymerase proteins and a nuclear signal in polymerase PB2[J]. Embo J, 1986, 5(9), 2371-6.
34. Jongkon, N., Mokmak, W., Chuakheaw, D., Shaw, P.J., Tongsima, S. and Sangma, C. Prediction of avian influenza A binding preference to human receptor using conformational analysis of receptor bound to hemagglutinin[J]. BMC Genomics, 2009, 10 Suppl 3, S24.
35. Jorba, N., Juarez, S., Torreira, E., Gastaminza, P., Zamarreno, N., Albar, J.P. and Ortin, J. Analysis of the interaction of influenza virus polymerase complex with human cell factors. [J] Proteomics, 2008, 8(10), 2077-88.
36. Kalthoff, D., Globig, A. and Beer, M. (Highly pathogenic) avian influenza as a zoonotic agent[J]. Vet Microbiol, 140(3-4), 237-45.
37. Kawaguchi, A. and Nagata, K. De novo replication of the influenza virus RNA genome is regulated by DNA replicative helicase, MCM[J]. Embo J, 2007, 26(21), 4566-75.
38. Kawaoka, Y., Chambers, T.M., Sladen, W.L. and Webster, R.G. Is the gene pool of influenza viruses in shorebirds and gulls different from that in wild ducks? [J] Virology, 1988,163(1), 247-50.
39. Klenk, H.D. and Rott, R. The molecular biology of influenza virus pathogenicity[J]. Adv Virus Res, 1988, 34, 247-81.
40. Leahy, M.B., Zecchin, G. and Brownlee, G.G. Differential activation of influenza A virus endonuclease activity is dependent on multiple sequence differences between the virion RNA and cRNA promoters[J]. J Virol, 2002, 76(4), 2019-23.
41. Lee, Y.J., Shin, J.Y., Song, M.S., Lee, Y.M., Choi, J.G., Lee, E.K., Jeong, O.M., Sung, H.W., Kim, J.H., Kwon, Y.K., Kwon, J.H., Kim, C.J., Webby, R.J., Webster, R.G. and Choi, Y.K. Continuing evolution of H9 influenza viruses in Korean poultry[J]. Virology, 2007, 359(2), 313-23.
42. Li, Z., Chen, H., Jiao, P., Deng, G., Tian, G., Li, Y., Hoffmann, E., Webster, R.G., Matsuoka, Y. and Yu, K. Molecular basis of replication of duck H5N1 influenza viruses in a mammalian mouse model[J]. J Virol, 2005, 79(18), 12058-64.
43. Matrosovich, M., Stech, J. and Klenk, H.D. Influenza receptors, polymerase and host range[J]. Rev Sci Tech, 2009, 28(1), 203-17.
44. Mayer, D., Molawi, K., Martinez-Sobrido, L., Ghanem, A., Thomas, S., Baginsky, S., Grossmann, J., Garcia-Sastre, A. and Schwemmle, M. Identification of cellular interaction partners of the influenza virus ribonucleoprotein complex and polymerase complex using proteomic-based approaches[J]. J Proteome Res, 2007, 6(2), 672-82.
45. Momose, F., Naito, T., Yano, K., Sugimoto, S., Morikawa, Y. and Nagata, K. Identification of Hsp90 as a stimulatory host factor involved in influenza virus RNA synthesis[J]. J Biol Chem, 2002, 277(47), 45306-14.
46. Nagata, K., Kawaguchi, A. and Naito, T. Host factors for replication and transcription of the influenza virus genome[J]. Rev Med Virol, 2008,247-60
47. Nakagawa, Y., Kimura, N., Toyoda, T., Mizumoto, K., Ishihama, A., Oda, K. and Nakada, S. The RNA polymerase PB2 subunit is not required for replication of the influenza virus genome but is involved in capped mRNA synthesis[J]. J Virol, 1995, 69(2), 728-33.
48. Neumann, G. and Kawaoka, Y. Host range restriction and pathogenicity in the context of influenza pandemic[J]. Emerg Infect Dis, 2006, 12(6), 881-6.
49. Ng, S.S., Li, O.T., Cheung, T.K., Malik Peiris, J.S. and Poon, L.L. Heterologous influenza vRNA segments with identical non-coding sequences stimulate viral RNA replication in trans[J]. Virol J ,2008,5, 2.
50. Nieto, A., de la Luna, S., Barcena, J., Portela, A. and Ortin, J. Complex structure of the nuclear translocation signal of influenza virus polymerase PA subunit[J]. J Gen Virol, 1994, 75(Pt1), 29-36.
51. Ohtsu, Y., Honda, Y., Sakata, Y., Kato, H. and Toyoda, T. Fine mapping of the subunit binding sites of influenza virus RNA polymerase[J]. Microbiol Immunol, 2002, 46(3), 167-75.
52. Olsen, B., Munster, V.J., Wallensten, A., Waldenstrom, J., Osterhaus, A.D. and Fouchier, R.A. Global patterns of influenza a virus in wild birds[J]. Science, 2006, 312(5772), 384-8.
53. Perez-Gonzalez, A., Rodriguez, A., Huarte, M., Salanueva, I.J. and Nieto, A. hCLE/CGI-99, a human protein that interacts with the influenza virus polymerase, is a mRNA transcription modulator[J]. J Mol Biol, 2006,362(5), 887-900.
54. Perez, D.R. and Donis, R.O. A 48-amino-acid region of influenza A virus PB1 protein is sufficient for complex formation with PA[J]. J Virol, 1995, 69(11), 6932-9.
55. Perez, D.R. and Donis, R.O. Functional analysis of PA binding by influenza a virus PB1: effects on polymerase activity and viral infectivity[J]. J Virol, 2001, 75(17), 8127-36.
56. Poole, E.L., Medcalf, L., Elton, D. and Digard, P. Evidence that the C-terminal PB2-binding region of the influenza A virus PB1 protein is a discrete alpha-helical domain[J]. FEBS Lett, 2007, 581(27), 5300-6.
57. Puig, O., Caspary, F., Rigaut, G., Rutz, B., Bouveret, E., Bragado-Nilsson, E., Wilm, M. and Seraphin, B. The tandem affinity purification (TAP) method: a general procedure of protein complex purification[J]. Methods, 2001, 24(3), 218-29.
58. Ray, K., Potdar, V.A., Cherian, S.S., Pawar, S.D., Jadhav, S.M., Waregaonkar, S.R., Joshi, A.A. and Mishra, A.C. Characterization of the complete genome of influenza A (H5N1) virus isolated during the 2006 outbreak in poultry in India[J]. Virus Genes, 2008, 36(2), 345-53.
59. Regan, J.E. (2005) Influenza A Virus A Role For The RNA Polymerase In Viral Particle Assembly.
60. Reperant, L.A., Rimmelzwaan, G.F. and Kuiken, T. Avian influenza viruses in mammals[J]. Rev Sci Tech, 2009, 28(1), 137-59.
61. Rigaut, G., Shevchenko, A., Rutz, B., Wilm, M., Mann, M. and Seraphin, B. A generic protein purification method for protein complex characterization and proteome exploration[J]. Nat Biotechnol, 1999, 17(10), 1030-2.
62. Robertson, J.S. 5' and 3' terminal nucleotide sequences of the RNA genome segments of influenza virus[J]. Nucleic Acids Res, 1979, 6(12), 3745-57.
63. Rott, R. The pathogenic determinant of influenza virus[J]. Vet Microbiol, 1992, 33(1-4), 303-10.
64. Slemons, R.D., Johnson, D.C., Osborn, J.S. and Hayes, F. Type-A influenza viruses isolated from wild free-flying ducks in California[J]. Avian Dis, 1974, 18(1), 119-24.
65. Subbarao, E.K., London, W. and Murphy, B.R. A single amino acid in the PB2 gene of influenza A virus is a determinant of host range[J]. J Virol, 1993, 67(4), 1761-4.
66. Snyder, M.H., Buckler-White, A.J., London, W.T., Tierney, E.L. and Murphy, B.R. The avian influenza virus nucleoprotein gene and a specific constellation of avian and human virus polymerase genes each specify attenuation of avian-human influenza A/Pintail/79 reassortant viruses for monkeys[J]. J Virol, 1987, 61(9), 2857-63.
67. Tarendeau, F., Boudet, J., Guilligay, D., Mas, P.J., Bougault, C.M., Boulo, S., Baudin, F., Ruigrok, R.W., Daigle, N., Ellenberg, J., Cusack, S., Simorre, J.P. and Hart, D.J. Structure and nuclear import function of the C-terminal domain of influenza virus polymerase PB2 subunit[J]. Nat Struct Mol Biol, 2007, 14(3), 229-33.
68. Vazquez de Aldana CR, Marton MJ, Hinnebusch AG. GCN20, a novel ATP binding cassette protein, and GCN1 reside in a complex that mediates activation of the eIF-2 alpha kinase GCN2 in amino acid-starved cells[J].EMBO J. 1995, 14(13), 3184-99.
69. Verdugo, C., Cardona, C.J. and Carpenter, T.E. Simulation of an early warning system using sentinel birds to detect a change of a low pathogenic avian influenza virus (LPAIV) to high pathogenic avian influenza virus (HPAIV) [J]. Prev Vet Med, 2009, 88(2), 109-19.
70. Vreede, F.T., Jung, T.E. and Brownlee, G.G. Model suggesting that replication of influenza virus is regulated by stabilization of replicative intermediates[J]. J Virol, 2004, 78(17), 9568-72.
71. Webster, R.G., Hinshaw, V.S., Bean, W.J. and Sriram, G. Influenza viruses: transmission between species[J]. Philos Trans R Soc Lond B Biol Sci, 1980, 288(1029), 439-47.
72. Yao, Y., Mingay, L.J., McCauley, J.W. and Barclay, W.S. Sequences in influenza A virus PB2 protein that determine productive infection for an avian influenza virus in mouse and human cell lines[J]. J Viro, 2001,l 75(11), 5410-5.
73. Zamarin, D., Garcia-Sastre, A., Xiao, X., Wang, R. and Palese, P. Influenza virus PB1-F2 protein induces cell death through mitochondrial ANT3 and VDAC1[J]. PLoS Pathog, 2005, 1(1), e4.
2.关薇,王建,贺福初.大规模蛋白质相互作用研究方法进展[J].生命科学,2006,18(5),507-511.
3.葛叶,邓国华,李雪平,陈化兰.流感病毒PB2亚单位抗血清的制备[J].中国预防兽医学报,已接受.
4.葛叶,邓国华,李雪平,陈化兰.流感病毒聚合酶PB1-1蛋白的表达与单克隆抗体的制备[J].中国预防兽医学报,2009,(9),726-729.
5.葛叶,邓国华,李雪平,高玉伟,姚秋成,唐艳玲,陈化兰.流感病毒聚合酶PA亚单位抗血清的制备[J].中国预防兽医学报,2009,(4),314-317.
6.童光志,王云峰.动物基因工程疫苗原理与方法[M].北京:化学工业出版社,2009,30-40
7.王海波,安学丽,张艳贞,王爱丽,李巧云,晏月明.蛋白质相互作用研究方法及其应用[J].生物技术通报,2006年增刊,167-170.
8.张成裕.免疫增强剂在动物医学上的研究进展及应用[J].福建畜牧兽医,2004(1),11-12.
9. Alexander, D.J. and Brown, I.H. Recent zoonoses caused by influenza A viruses[J]. Rev Sci Tech ,2000,19(1), 197-225.
10. Area, E., Martin-Benito, J., Gastaminza, P., Torreira, E., Valpuesta, J.M., Carrascosa, J.L. and Ortin, J. 3D structure of the influenza virus polymerase complex: localization of subunit domains[J]. Proc Natl Acad Sci U S A ,2004,101(1), 308-13.
11. Basler, C.F. and Aguilar, P.V. Progress in identifying virulence determinants of the 1918 H1N1 and the Southeast Asian H5N1 influenza A viruses[J]. Antiviral Res ,2008,79(3) 166-78.
12. Biswas, S.K. and Nayak, D.P. Mutational analysis of the conserved motifs of influenza A virus polymerase basic protein 1[J]. J Virol 6,1994,8(3), 1819-26.
13. Boulo, S., Akarsu, H., Ruigrok, R.W. and Baudin, F. Nuclear traffic of influenza virus proteins and ribonucleoprotein complexes[J]. Virus Res ,2007,124(1-2), 12-21.
14. Burgui, I., Yanguez, E., Sonenberg, N. and Nieto, A. Influenza virus mRNA translation revisited: is the eIF4E cap-binding factor required for viral mRNA translation? [J] J Virol ,2007,81(22), 12427-38.
15. Cardona, C.J., Xing, Z., Sandrock, C.E. and Davis, C.E. Avian influenza in birds and mammals[J]. Comp Immunol Microbiol Infect Dis ,2009,32(4), 255-73.
16. Chan, A.Y., Vreede, F.T., Smith, M., Engelhardt, O.G. and Fodor, E. Influenza virus inhibits RNA polymerase II elongation[J]. Virology ,2006,351(1), 210-7.
17. Deng, T., Engelhardt, O.G., Thomas, B., Akoulitchev, A.V., Brownlee, G.G. and Fodor, E. Role of ran binding protein 5 in nuclear import and assembly of the influenza virus RNA polymerase complex[J]. J Virol ,2006,80(24), 11911-9.
18. Desselberger, U., Racaniello, V.R., Zazra, J.J. and Palese, P. , The 3' and 5'-terminal sequences of influenza A, B and C virus RNA segments are highly conserved and show partial inverted complementarity[J]. Gene,1980, 8(3), 315-28.
19. Engelhardt, O.G., Smith, M. and Fodor, E. Association of the influenza A virus RNA-dependent RNA polymerase with cellular RNA polymerase II[J]. J Virol ,2005,79(9), 5812-8.
20. Fechter, P., Mingay, L., Sharps, J., Chambers, A., Fodor, E. and Brownlee, G.G. Two aromatic residues in the PB2 subunit of influenza A RNA polymerase are crucial for cap binding[J]. J Biol Chem ,2003,278(22), 20381-8.
21. Flick, R. and Hobom, G. Interaction of influenza virus polymerase with viral RNA in the 'corkscrew' conformation[J]. J Gen Virol ,1999,80 (Pt 10), 2565-72.
22. Fodor, E., Pritlove, D.C. and Brownlee, G.G. Characterization of the RNA-fork model of virion RNA in the initiation of transcription in influenza A virus[J]. J Virol ,1995,69(7), 4012-9.
23. Fujii, K., Fujii, Y., Noda, T., Muramoto, Y., Watanabe, T., Takada, A., Goto, H., Horimoto, T. and Kawaoka, Y. Importance of both the coding and the segment-specific noncoding regions of the influenza A virus NS segment for its efficient incorporation into virions[J]. J Virol, 2005, 79(6), 3766-74.
24. Gabriel, G., Abram, M., Keiner, B., Wagner, R., Klenk, H.D. and Stech, J. Differential polymerase activity in avian and mammalian cells determines host range of influenza virus[J]. J Virol, 2007, 81(17), 9601-4.
25. Gabriel, G., Herwig, A. and Klenk, H.D. Interaction of polymerase subunit PB2 and NP with importin alpha1 is a determinant of host range of influenza A virus[J]. PLoS Pathog, 2008, 4(2), e11.
26. Hara, K., Schmidt, F.I., Crow, M. and Brownlee, G.G. Amino acid residues in the N-terminal region of the PA subunit of influenza A virus RNA polymerase play a critical role in protein stability, endonuclease activity, cap binding, and virion RNA promoter binding[J]. J Virol, 2006,80(16), 7789-98.
27. Honda, A., Mizumoto, K. and Ishihama, A. Two separate sequences of PB2 subunit constitute the RNA cap-binding site of influenza virus RNA polymerase[J]. Genes Cells ,1999,4(8), 475-85.
28. Hossain, M.J., Mori, I., Dong, L., Liu, B. and Kimura, Y. Fetal calf serum inhibits virus genome expression in Madin-Darby canine kidney cells persistently infected with influenza A virus[J]. Med Microbiol Immunol, 2008, 197(1), 21-7.
29. Hsu, M.T., Parvin, J.D., Gupta, S., Krystal, M. and Palese, P. Genomic RNAs of influenza viruses are held in a circular conformation in virions and in infected cells by a terminal panhandle[J]. Proc Natl Acad Sci U S A, 1987, 84(22), 8140-4.
30. Huarte, M., Sanz-Ezquerro, J.J., Roncal, F., Ortin, J. and Nieto, A. PA subunit from influenza virus polymerase complex interacts with a cellular protein with homology to a family of transcriptional activators[J]. J Virol, 2001, 75(18), 8597-604.
31. Hynes RO. The extracellular matrix: not just pretty fibrils[J].Science, 2009, 326(5957),1216-9.
32. Ito, T. and Kawaoka, Y. Host-range barrier of influenza A viruses[J]. Vet Microbiol, 2000, 74(1-2), 71-5.
33. Jones, I.M., Reay, P.A. and Philpott, K.L. Nuclear location of all three influenza polymerase proteins and a nuclear signal in polymerase PB2[J]. Embo J, 1986, 5(9), 2371-6.
34. Jongkon, N., Mokmak, W., Chuakheaw, D., Shaw, P.J., Tongsima, S. and Sangma, C. Prediction of avian influenza A binding preference to human receptor using conformational analysis of receptor bound to hemagglutinin[J]. BMC Genomics, 2009, 10 Suppl 3, S24.
35. Jorba, N., Juarez, S., Torreira, E., Gastaminza, P., Zamarreno, N., Albar, J.P. and Ortin, J. Analysis of the interaction of influenza virus polymerase complex with human cell factors. [J] Proteomics, 2008, 8(10), 2077-88.
36. Kalthoff, D., Globig, A. and Beer, M. (Highly pathogenic) avian influenza as a zoonotic agent[J]. Vet Microbiol, 140(3-4), 237-45.
37. Kawaguchi, A. and Nagata, K. De novo replication of the influenza virus RNA genome is regulated by DNA replicative helicase, MCM[J]. Embo J, 2007, 26(21), 4566-75.
38. Kawaoka, Y., Chambers, T.M., Sladen, W.L. and Webster, R.G. Is the gene pool of influenza viruses in shorebirds and gulls different from that in wild ducks? [J] Virology, 1988,163(1), 247-50.
39. Klenk, H.D. and Rott, R. The molecular biology of influenza virus pathogenicity[J]. Adv Virus Res, 1988, 34, 247-81.
40. Leahy, M.B., Zecchin, G. and Brownlee, G.G. Differential activation of influenza A virus endonuclease activity is dependent on multiple sequence differences between the virion RNA and cRNA promoters[J]. J Virol, 2002, 76(4), 2019-23.
41. Lee, Y.J., Shin, J.Y., Song, M.S., Lee, Y.M., Choi, J.G., Lee, E.K., Jeong, O.M., Sung, H.W., Kim, J.H., Kwon, Y.K., Kwon, J.H., Kim, C.J., Webby, R.J., Webster, R.G. and Choi, Y.K. Continuing evolution of H9 influenza viruses in Korean poultry[J]. Virology, 2007, 359(2), 313-23.
42. Li, Z., Chen, H., Jiao, P., Deng, G., Tian, G., Li, Y., Hoffmann, E., Webster, R.G., Matsuoka, Y. and Yu, K. Molecular basis of replication of duck H5N1 influenza viruses in a mammalian mouse model[J]. J Virol, 2005, 79(18), 12058-64.
43. Matrosovich, M., Stech, J. and Klenk, H.D. Influenza receptors, polymerase and host range[J]. Rev Sci Tech, 2009, 28(1), 203-17.
44. Mayer, D., Molawi, K., Martinez-Sobrido, L., Ghanem, A., Thomas, S., Baginsky, S., Grossmann, J., Garcia-Sastre, A. and Schwemmle, M. Identification of cellular interaction partners of the influenza virus ribonucleoprotein complex and polymerase complex using proteomic-based approaches[J]. J Proteome Res, 2007, 6(2), 672-82.
45. Momose, F., Naito, T., Yano, K., Sugimoto, S., Morikawa, Y. and Nagata, K. Identification of Hsp90 as a stimulatory host factor involved in influenza virus RNA synthesis[J]. J Biol Chem, 2002, 277(47), 45306-14.
46. Nagata, K., Kawaguchi, A. and Naito, T. Host factors for replication and transcription of the influenza virus genome[J]. Rev Med Virol, 2008,247-60
47. Nakagawa, Y., Kimura, N., Toyoda, T., Mizumoto, K., Ishihama, A., Oda, K. and Nakada, S. The RNA polymerase PB2 subunit is not required for replication of the influenza virus genome but is involved in capped mRNA synthesis[J]. J Virol, 1995, 69(2), 728-33.
48. Neumann, G. and Kawaoka, Y. Host range restriction and pathogenicity in the context of influenza pandemic[J]. Emerg Infect Dis, 2006, 12(6), 881-6.
49. Ng, S.S., Li, O.T., Cheung, T.K., Malik Peiris, J.S. and Poon, L.L. Heterologous influenza vRNA segments with identical non-coding sequences stimulate viral RNA replication in trans[J]. Virol J ,2008,5, 2.
50. Nieto, A., de la Luna, S., Barcena, J., Portela, A. and Ortin, J. Complex structure of the nuclear translocation signal of influenza virus polymerase PA subunit[J]. J Gen Virol, 1994, 75(Pt1), 29-36.
51. Ohtsu, Y., Honda, Y., Sakata, Y., Kato, H. and Toyoda, T. Fine mapping of the subunit binding sites of influenza virus RNA polymerase[J]. Microbiol Immunol, 2002, 46(3), 167-75.
52. Olsen, B., Munster, V.J., Wallensten, A., Waldenstrom, J., Osterhaus, A.D. and Fouchier, R.A. Global patterns of influenza a virus in wild birds[J]. Science, 2006, 312(5772), 384-8.
53. Perez-Gonzalez, A., Rodriguez, A., Huarte, M., Salanueva, I.J. and Nieto, A. hCLE/CGI-99, a human protein that interacts with the influenza virus polymerase, is a mRNA transcription modulator[J]. J Mol Biol, 2006,362(5), 887-900.
54. Perez, D.R. and Donis, R.O. A 48-amino-acid region of influenza A virus PB1 protein is sufficient for complex formation with PA[J]. J Virol, 1995, 69(11), 6932-9.
55. Perez, D.R. and Donis, R.O. Functional analysis of PA binding by influenza a virus PB1: effects on polymerase activity and viral infectivity[J]. J Virol, 2001, 75(17), 8127-36.
56. Poole, E.L., Medcalf, L., Elton, D. and Digard, P. Evidence that the C-terminal PB2-binding region of the influenza A virus PB1 protein is a discrete alpha-helical domain[J]. FEBS Lett, 2007, 581(27), 5300-6.
57. Puig, O., Caspary, F., Rigaut, G., Rutz, B., Bouveret, E., Bragado-Nilsson, E., Wilm, M. and Seraphin, B. The tandem affinity purification (TAP) method: a general procedure of protein complex purification[J]. Methods, 2001, 24(3), 218-29.
58. Ray, K., Potdar, V.A., Cherian, S.S., Pawar, S.D., Jadhav, S.M., Waregaonkar, S.R., Joshi, A.A. and Mishra, A.C. Characterization of the complete genome of influenza A (H5N1) virus isolated during the 2006 outbreak in poultry in India[J]. Virus Genes, 2008, 36(2), 345-53.
59. Regan, J.E. (2005) Influenza A Virus A Role For The RNA Polymerase In Viral Particle Assembly.
60. Reperant, L.A., Rimmelzwaan, G.F. and Kuiken, T. Avian influenza viruses in mammals[J]. Rev Sci Tech, 2009, 28(1), 137-59.
61. Rigaut, G., Shevchenko, A., Rutz, B., Wilm, M., Mann, M. and Seraphin, B. A generic protein purification method for protein complex characterization and proteome exploration[J]. Nat Biotechnol, 1999, 17(10), 1030-2.
62. Robertson, J.S. 5' and 3' terminal nucleotide sequences of the RNA genome segments of influenza virus[J]. Nucleic Acids Res, 1979, 6(12), 3745-57.
63. Rott, R. The pathogenic determinant of influenza virus[J]. Vet Microbiol, 1992, 33(1-4), 303-10.
64. Slemons, R.D., Johnson, D.C., Osborn, J.S. and Hayes, F. Type-A influenza viruses isolated from wild free-flying ducks in California[J]. Avian Dis, 1974, 18(1), 119-24.
65. Subbarao, E.K., London, W. and Murphy, B.R. A single amino acid in the PB2 gene of influenza A virus is a determinant of host range[J]. J Virol, 1993, 67(4), 1761-4.
66. Snyder, M.H., Buckler-White, A.J., London, W.T., Tierney, E.L. and Murphy, B.R. The avian influenza virus nucleoprotein gene and a specific constellation of avian and human virus polymerase genes each specify attenuation of avian-human influenza A/Pintail/79 reassortant viruses for monkeys[J]. J Virol, 1987, 61(9), 2857-63.
67. Tarendeau, F., Boudet, J., Guilligay, D., Mas, P.J., Bougault, C.M., Boulo, S., Baudin, F., Ruigrok, R.W., Daigle, N., Ellenberg, J., Cusack, S., Simorre, J.P. and Hart, D.J. Structure and nuclear import function of the C-terminal domain of influenza virus polymerase PB2 subunit[J]. Nat Struct Mol Biol, 2007, 14(3), 229-33.
68. Vazquez de Aldana CR, Marton MJ, Hinnebusch AG. GCN20, a novel ATP binding cassette protein, and GCN1 reside in a complex that mediates activation of the eIF-2 alpha kinase GCN2 in amino acid-starved cells[J].EMBO J. 1995, 14(13), 3184-99.
69. Verdugo, C., Cardona, C.J. and Carpenter, T.E. Simulation of an early warning system using sentinel birds to detect a change of a low pathogenic avian influenza virus (LPAIV) to high pathogenic avian influenza virus (HPAIV) [J]. Prev Vet Med, 2009, 88(2), 109-19.
70. Vreede, F.T., Jung, T.E. and Brownlee, G.G. Model suggesting that replication of influenza virus is regulated by stabilization of replicative intermediates[J]. J Virol, 2004, 78(17), 9568-72.
71. Webster, R.G., Hinshaw, V.S., Bean, W.J. and Sriram, G. Influenza viruses: transmission between species[J]. Philos Trans R Soc Lond B Biol Sci, 1980, 288(1029), 439-47.
72. Yao, Y., Mingay, L.J., McCauley, J.W. and Barclay, W.S. Sequences in influenza A virus PB2 protein that determine productive infection for an avian influenza virus in mouse and human cell lines[J]. J Viro, 2001,l 75(11), 5410-5.
73. Zamarin, D., Garcia-Sastre, A., Xiao, X., Wang, R. and Palese, P. Influenza virus PB1-F2 protein induces cell death through mitochondrial ANT3 and VDAC1[J]. PLoS Pathog, 2005, 1(1), e4.