Inhibition of Apoptosis and NF-κB Activation by Vaccinia Protein N1 Occur via Distinct Binding Surfaces and Make Different Contributions to Virulence

English version České info

Vaccinia virus (VACV) protein N1 is an intracellular virulence factor and belongs to a family of VACV B-cell lymphoma (Bcl)-2-like proteins whose members inhibit apoptosis or activation of pro-inflammatory transcription factors, such as interferon (IFN) regulatory factor-3 (IRF-3) and nuclear factor-κB (NF-κB). Unusually, N1 inhibits both apoptosis and NF-κB activation. To understand how N1 exerts these different functions, we have mutated residues in the Bcl-2-like surface groove and at the interface used to form N1 homodimers. Mutagenesis of the surface groove abolished only the N1 anti-apoptotic activity and protein crystallography showed these mutants differed from wild-type N1 only at the site of mutation. Conversely, mutagenesis of the dimer interface converted N1 to a monomer and affected only inhibition of NF-κB activation. Collectively, these data show that N1 inhibits pro-inflammatory and pro-apoptotic signalling using independent surfaces of the protein. To determine the relative contribution of each activity to virus virulence, mutant N1 alleles were introduced into a VACV strain lacking N1 and the virulence of these viruses was analysed after intradermal and intranasal inoculation in mice. In both models, VACV containing a mutant N1 unable to inhibit apoptosis had similar virulence to wild-type virus, whereas VACV containing a mutant N1 impaired for NF-κB inhibition induced an attenuated infection similar to that of the N1-deleted virus. This indicates that anti-apoptotic activity of N1 does not drive virulence in these in vivo models, and highlights the importance of pro-inflammatory signalling in the immune response against viral infections.

Vyšlo v časopise: Inhibition of Apoptosis and NF-κB Activation by Vaccinia Protein N1 Occur via Distinct Binding Surfaces and Make Different Contributions to Virulence. PLoS Pathog 7(12): e32767. doi:10.1371/journal.ppat.1002430
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.ppat.1002430

Souhrn

Zdroje

1. MohamedMRMcFaddenG 2009 NFkB inhibitors: strategies from poxviruses. Cell Cycle 8 3125 3132

2. SeetBTJohnstonJBBrunettiCRBarrettJWEverettH 2003 Poxviruses and immune evasion. Annu Rev Immunol 21 377 423

3. BartlettNSymonsJATscharkeDCSmithGL 2002 The vaccinia virus N1L protein is an intracellular homodimer that promotes virulence. J Gen Virol 83 1965 1976

4. JacobsNBartlettNWClarkRHSmithGL 2008 Vaccinia virus lacking the Bcl-2-like protein N1 induces a stronger natural killer cell response to infection. J Gen Virol 89 2877 2881

5. KotwalGJHuginAWMossB 1989 Mapping and insertional mutagenesis of a vaccinia virus gene encoding a 13,800-Da secreted protein. Virology 171 579 587

6. AoyagiMZhaiDJinCAleshinAEStecB 2007 Vaccinia virus N1L protein resembles a B cell lymphoma-2 (Bcl-2) family protein. Protein Sci 16 118 124

7. GrahamSCBaharMWCooraySChenRAWhalenDM 2008 Vaccinia virus proteins A52 and B14 Share a Bcl-2-like fold but have evolved to inhibit NF-kappaB rather than apoptosis. PLoS Pathog 4 e1000128

8. CooraySBaharMWAbresciaNGMcVeyCEBartlettNW 2007 Functional and structural studies of the vaccinia virus virulence factor N1 reveal a Bcl-2-like anti-apoptotic protein. J Gen Virol 88 1656 1666

9. ChenRARyzhakovGCooraySRandowFSmithGL 2008 Inhibition of IkappaB kinase by vaccinia virus virulence factor B14. PLoS Pathog 4 e22

10. KalverdaAPThompsonGSVogelASchroderMBowieAG 2009 Poxvirus K7 protein adopts a Bcl-2 fold: biochemical mapping of its interactions with human DEAD box RNA helicase DDX3. J Mol Biol 385 843 853

11. BenfieldCTMansurDSMcCoyLEFergusonBJBaharMW 2011 Mapping the IkappaB kinase beta (IKKbeta)-binding interface of the B14 protein, a vaccinia virus inhibitor of IKKbeta-mediated activation of nuclear factor kappaB. J Biol Chem 286 20727 20735

12. GonzalezJMEstebanM 2010 A poxvirus Bcl-2-like gene family involved in regulation of host immune response: sequence similarity and evolutionary history. Virol J 7 59

13. DiPernaGStackJBowieAGBoydAKotwalG 2004 Poxvirus protein N1L targets the I-kappaB kinase complex, inhibits signaling to NF-kappaB by the tumor necrosis factor superfamily of receptors, and inhibits NF-kappaB and IRF3 signaling by toll-like receptors. J Biol Chem 279 36570 36578

14. HaydenMSGhoshS 2008 Shared principles in NF-kappaB signaling. Cell 132 344 362

15. HaydenMSWestAPGhoshS 2006 NF-kappaB and the immune response. Oncogene 25 6758 6780

16. AkiraSUematsuSTakeuchiO 2006 Pathogen recognition and innate immunity. Cell 124 783 801

17. TaitSWGreenDR 2010 Mitochondria and cell death: outer membrane permeabilization and beyond. Nat Rev Mol Cell Biol 11 621 632

18. LamkanfiMDixitVM 2010 Manipulation of host cell death pathways during microbial infections. Cell Host Microbe 8 44 54

19. GalluzziLBrennerCMorselliETouatZKroemerG 2008 Viral control of mitochondrial apoptosis. PLoS Pathog 4 e1000018

20. HuangQPetrosAMVirginHWFesikSWOlejniczakET 2002 Solution structure of a Bcl-2 homolog from Kaposi sarcoma virus. Proc Natl Acad Sci U S A 99 3428 3433

21. SaridRSatoTBohenzkyRARussoJJChangY 1997 Kaposi's sarcoma-associated herpesvirus encodes a functional bcl-2 homologue. Nat Med 3 293 298

22. HuangQPetrosAMVirginHWFesikSWOlejniczakET 2003 Solution structure of the BHRF1 protein from Epstein-Barr virus, a homolog of human Bcl-2. J Mol Biol 332 1123 1130

23. HendersonSHuenDRoweMDawsonCJohnsonG 1993 Epstein-Barr virus-coded BHRF1 protein, a viral homologue of Bcl-2, protects human B cells from programmed cell death. Proc Natl Acad Sci U S A 90 8479 8483

24. KvansakulMWeiAHFletcherJIWillisSNChenL 2010 Structural basis for apoptosis inhibition by Epstein-Barr virus BHRF1. PLoS Pathog 6 e1001236

25. LohJHuangQPetrosAMNettesheimDvan DykLF 2005 A surface groove essential for viral Bcl-2 function during chronic infection in vivo. PLoS Pathog 1 e10

26. VirginHWtLatreillePWamsleyPHallsworthKWeckKE 1997 Complete sequence and genomic analysis of murine gammaherpesvirus 68. J Virol 71 5894 5904

27. JohnstonJBMcFaddenG 2003 Poxvirus immunomodulatory strategies: current perspectives. J Virol 77 6093 6100

28. KvansakulMvan DelftMFLeeEFGulbisJMFairlieWD 2007 A structural viral mimic of prosurvival Bcl-2: a pivotal role for sequestering proapoptotic Bax and Bak. Mol Cell 25 933 942

29. DouglasAECorbettKDBergerJMMcFaddenGHandelTM 2007 Structure of M11L: A myxoma virus structural homolog of the apoptosis inhibitor, Bcl-2. Protein Sci 16 695 703

30. GrahamKAOpgenorthAUptonCMcFaddenG 1992 Myxoma virus M11L ORF encodes a protein for which cell surface localization is critical in manifestation of viral virulence. Virology 191 112 124

31. CampbellSHazesBKvansakulMColmanPBarryM 2010 Vaccinia virus F1L interacts with Bak using highly divergent Bcl-2 homology domains and replaces the function of Mcl-1. J Biol Chem 285 4695 4708

32. PostigoACrossJRDownwardJWayM 2006 Interaction of F1L with the BH3 domain of Bak is responsible for inhibiting vaccinia-induced apoptosis. Cell Death Differ 13 1651 1662

33. WasilenkoSTBanadygaLBondDBarryM 2005 The vaccinia virus F1L protein interacts with the proapoptotic protein Bak and inhibits Bak activation. J Virol 79 14031 14043

34. WasilenkoSTStewartTLMeyersAFBarryM 2003 Vaccinia virus encodes a previously uncharacterized mitochondrial-associated inhibitor of apoptosis. Proc Natl Acad Sci U S A 100 14345 14350

35. KvansakulMYangHFairlieWDCzabotarPEFischerSF 2008 Vaccinia virus anti-apoptotic F1L is a novel Bcl-2-like domain-swapped dimer that binds a highly selective subset of BH3-containing death ligands. Cell Death Differ 15 1564 1571

36. ChipukJEMoldoveanuTLlambiFParsonsMJGreenDR 2010 The BCL-2 family reunion. Mol Cell 37 299 310

37. YouleRJStrasserA 2008 The BCL-2 protein family: opposing activities that mediate cell death. Nat Rev Mol Cell Biol 9 47 59

38. GiamMHuangDCBouilletP 2008 BH3-only proteins and their roles in programmed cell death. Oncogene 27 Suppl 1 S128 136

39. OdaSSchroderMKhanAR 2009 Structural basis for targeting of human RNA helicase DDX3 by poxvirus protein K7. Structure 17 1528 1537

40. MaraniMTenevTHancockDDownwardJLemoineNR 2002 Identification of novel isoforms of the BH3 domain protein Bim which directly activate Bax to trigger apoptosis. Mol Cell Biol 22 3577 3589

41. WillisSNFletcherJIKaufmannTvan DelftMFChenL 2007 Apoptosis initiated when BH3 ligands engage multiple Bcl-2 homologs, not Bax or Bak. Science 315 856 859

42. WillisSNAdamsJM 2005 Life in the balance: how BH3-only proteins induce apoptosis. Curr Opin Cell Biol 17 617 625

43. LovellJFBillenLPBindnerSShamas-DinAFradinC 2008 Membrane binding by tBid initiates an ordered series of events culminating in membrane permeabilization by Bax. Cell 135 1074 1084

44. LeberBLinJAndrewsDW 2007 Embedded together: the life and death consequences of interaction of the Bcl-2 family with membranes. Apoptosis 12 897 911

45. WangGBarrettJWNazarianSHEverettHGaoX 2004 Myxoma virus M11L prevents apoptosis through constitutive interaction with Bak. J Virol 78 7097 7111

46. GavathiotisESuzukiMDavisMLPitterKBirdGH 2008 BAX activation is initiated at a novel interaction site. Nature 455 1076 1081

47. MathewAO'BryanJMarshallWKotwalGJTerajimaM 2008 Robust intrapulmonary CD8 T cell responses and protection with an attenuated N1L deleted vaccinia virus. PLoS One 3 e3323

48. GratzMSSuezerYKremerMVolzAMajzoubM 2011 N1L is an ectromelia virus virulence factor and essential for in vivo spread upon respiratory infection. J Virol 85 3557 3569

49. GloecknerCJBoldtKSchumacherARoepmanRUeffingM 2007 A novel tandem affinity purification strategy for the efficient isolation and characterisation of native protein complexes. Proteomics 7 4228 4234

50. RandowFSaleJE 2006 Retroviral transduction of DT40. Subcell Biochem 40 383 386

51. ChenRAJacobsNSmithGL 2006 Vaccinia virus strain Western Reserve protein B14 is an intracellular virulence factor. J Gen Virol 87 1451 1458

52. ParkinsonJESmithGL 1994 Vaccinia virus gene A36R encodes a M(r) 43-50 K protein on the surface of extracellular enveloped virus. Virology 204 376 390

53. EmsleyPLohkampBScottWGCowtanK 2010 Features and development of Coot. Acta Crystallogr D Biol Crystallogr 66 486 501

54. SmartOSBrandlMFlensburgCKellerPPaciorekW 2008 Refinement with Local Structure Similarity Restraints (LSSR) Enables Exploitation of Information from Related Structures and Facilitates use of NCS. Abstr Annu Meet Am Crystallogr Assoc Abstract TP139 117

55. ChenVBArendall3rdWBHeaddJJKeedyDAImmorminoRM 2010 MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr D Biol Crystallogr 66 12 21

56. KrissinelEHenrickK 2004 Secondary-structure matching (SSM), a new tool for fast protein structure alignment in three dimensions. Acta Crystallogr D Biol Crystallogr 60 2256 2268

57. FalknerFGMossB 1990 Transient dominant selection of recombinant vaccinia viruses. J Virol 64 3108 3111

58. BoyleDBCouparBE 1988 A dominant selectable marker for the construction of recombinant poxviruses. Gene 65 123 128

59. KerrSMSmithGL 1991 Vaccinia virus DNA ligase is nonessential for virus replication: recovery of plasmids from virus-infected cells. Virology 180 625 632

60. MellonPParkerVGluzmanYManiatisT 1981 Identification of DNA sequences required for transcription of the human alpha 1-globin gene in a new SV40 host-vector system. Cell 27 279 288

61. Barrios-RodilesMBrownKROzdamarBBoseRLiuZ 2005 High-throughput mapping of a dynamic signaling network in mammalian cells. Science 307 1621 1625

62. AlcamiASmithGL 1992 A soluble receptor for interleukin-1 beta encoded by vaccinia virus: a novel mechanism of virus modulation of the host response to infection. Cell 71 153 167

63. TscharkeDCReadingPCSmithGL 2002 Dermal infection with vaccinia virus reveals roles for virus proteins not seen using other inoculation routes. J Gen Virol 83 1977 1986