Suppression of RNA Silencing by a Plant DNA Virus Satellite Requires a Host Calmodulin-Like Protein to Repress Expression

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In plants, RNA silencing plays a key role in antiviral defense. To counteract host defense, plant viruses encode viral suppressors of RNA silencing (VSRs) that target different effector molecules in the RNA silencing pathway. Evidence has shown that plants also encode endogenous suppressors of RNA silencing (ESRs) that function in proper regulation of RNA silencing. The possibility that these cellular proteins can be subverted by viruses to thwart host defense is intriguing but has not been fully explored. Here we report that the Nicotiana benthamiana calmodulin-like protein Nbrgs-CaM is required for the functions of the VSR βC1, the sole protein encoded by the DNA satellite associated with the geminivirus Tomato yellow leaf curl China virus (TYLCCNV). Nbrgs-CaM expression is up-regulated by the βC1. Transgenic plants over-expressing Nbrgs-CaM displayed developmental abnormities reminiscent of βC1-associated morphological alterations. Nbrgs-CaM suppressed RNA silencing in an Agrobacterium infiltration assay and, when over-expressed, blocked TYLCCNV-induced gene silencing. Genetic evidence showed that Nbrgs-CaM mediated the βC1 functions in silencing suppression and symptom modulation, and was required for efficient virus infection. Moreover, the tobacco and tomato orthologs of Nbrgs-CaM also possessed ESR activity, and were induced by betasatellite to promote virus infection in these Solanaceae hosts. We further demonstrated that βC1-induced Nbrgs-CaM suppressed the production of secondary siRNAs, likely through repressing RNA-DEPENDENT RNA POLYMERASE 6 (RDR6) expression. RDR6-deficient N. benthamiana plants were defective in antiviral response and were hypersensitive to TYLCCNV infection. More significantly, TYLCCNV could overcome host range restrictions to infect Arabidopsis thaliana when the plants carried a RDR6 mutation. These findings demonstrate a distinct mechanism of VSR for suppressing PTGS through usurpation of a host ESR, and highlight an essential role for RDR6 in RNA silencing defense response against geminivirus infection.

Vyšlo v časopise: Suppression of RNA Silencing by a Plant DNA Virus Satellite Requires a Host Calmodulin-Like Protein to Repress Expression. PLoS Pathog 10(2): e32767. doi:10.1371/journal.ppat.1003921
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.ppat.1003921

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Zdroje

1. DingSW (2010) RNA-based antiviral immunity. Nat Rev Immunol 10: 632–644.

2. LlaveC (2010) Virus-derived small interfering RNAs at the core of plant-virus interactions. Trends Plant Sci 15: 701–707.

3. DalmayT, HamiltonA, RuddS, AngellS, BaulcombeDC (2000) An RNA-dependent RNA polymerase gene in Arabidopsis is required for posttranscriptional gene silencing mediated by a transgene but not by a virus. Cell 101: 543–553.

4. MourrainP, BéclinC, ElmayanT, FeuerbachF, GodonC, et al. (2000) Arabidopsis SGS2 and SGS3 genes are required for posttranscriptional gene silencing and natural virus resistance. Cell 101: 533–542.

5. XieZ, FanB, ChenC, ChenZ (2001) An important role of an inducible RNA-dependent RNA polymerase in plant antiviral defense. Proc Natl Acad Sci U S A 98: 6516–6521.

6. Garcia-RuizH, TakedaA, ChapmanEJ, SullivanCM, FahlgrenN, et al. (2010) Arabidopsis RNA-dependent RNA polymerases and dicer-like proteins in antiviral defense and small interfering RNA biogenesis during Turnip mosaic virus infection. Plant Cell 22: 481–496.

7. MoissiardG, ParizottoEA, HimberC, VoinnetO (2007) Transitivity in Arabidopsis can be primed, requires the redundant action of the antiviral Dicer-like 4 and Dicer-like 2, and is compromised by viral-encoded suppressor proteins. RNA 13: 1268–1278.

8. QuF (2010) Antiviral role of plant-encoded RNA-dependent RNA polymerases revisited with deep sequencing of small interfering RNAs of virus origin. Mol Plant Microbe Interact 23: 1248–1252.

9. WangXB, WuQ, ItoT, CilloF, LiWX, et al. (2010) RNAi-mediated viral immunity requires amplification of virus-derived siRNAs in Arabidopsis thaliana. Proc Natl Acad Sci U S A 107: 484–489.

10. WangXB, JovelJ, UdompornP, WangY, WuQ, et al. (2011) The 21-nucleotide, but not 22-nucleotide, viral secondary small interfering RNAs direct potent antiviral defense by two cooperative argonautes in Arabidopsis thaliana. Plant Cell 23: 1625–1638.

11. Di SerioF, Martinez de AlbaAE, NavarroB, GiselA, FloresR (2010) RNA-dependent RNA polymerase 6 delays accumulation and precludes meristem invasion of a viroid that replicates in the nucleus. J Virol 84: 2477–2489.

12. QuF, YeX, HouG, SatoS, ClementeTE, et al. (2005) RDR6 has a broad-spectrum but temperature-dependent antiviral defense role in Nicotiana benthamiana. J Virol 79: 15209–15217.

13. SchwachF, VaistijFE, JonesL, BaulcombeDC (2005) An RNA-dependent RNA polymerase prevents meristem invasion by Potato virus X and is required for the activity but not the production of a systemic silencing signal. Plant Physiol 138: 1842–1852.

14. VaistijFE, JonesL (2009) Compromised virus-induced gene silencing in RDR6-deficient plants. Plant Physiol 149: 1399–1407.

15. BurgyanJ, HaveldaZ (2011) Viral suppressors of RNA silencing. Trends Plant Sci 16: 265–272.

16. WuQ, WangX, DingSW (2010) Viral suppressors of RNA-based viral immunity: host targets. Cell Host Microbe 8: 12–15.

17. ChaoJA, LeeJH, ChapadosBR, DeblerEW, SchneemannA, et al. (2005) Dual modes of RNA-silencing suppression by flock house virus protein B2. Nat Struct Mol Biol 12: 952–957.

18. ChenHY, YangJ, LinC, YuanYA (2008) Structural basis for RNA-silencing suppression by Tomato aspermy virus protein 2b. EMBO Rep 9: 754–760.

19. MeraiZ, KerenyiZ, KerteszS, MagnaM, LakatosL, et al. (2006) Double-stranded RNA binding may be a general plant RNA viral strategy to suppress RNA silencing. J Virol 80: 5747–5756.

20. VargasonJM, SzittyaG, BurgyanJ, HallTM (2003) Size selective recognition of siRNA by an RNA silencing suppressor. Cell 115: 799–811.

21. HaasG, AzevedoJ, MoissiardG, GeldreichA, HimberC, et al. (2008) Nuclear import of CaMV P6 is required for infection and suppression of the RNA silencing factor DRB4. EMBO J 27: 2102–2112.

22. GinerA, LakatosL, Garcia-ChapaM, Lopez-MoyaJJ, BurgyanJ (2010) Viral protein inhibits RISC activity by argonaute binding through conserved WG/GW motifs. PLoS Pathog 6: e1000996 doi: 10.1371/journal.ppat.1000996

23. ZhangX, YuanYR, PeiY, LinSS, TuschlT, et al. (2006) Cucumber mosaic virus-encoded 2b suppressor inhibits Arabidopsis ARGONAUTE1 cleavage activity to counter plant defense. Genes Dev 20: 3255–3268.

24. BortolamiolD, PazhouhandehM, MarroccoK, GenschikP, Ziegler-GraffV (2007) The Polerovirus F box protein P0 targets ARGONAUTE1 to suppress RNA silencing. Curr Biol 17: 1615–1621.

25. Csorba T, Lozsa R, Hutvagner G, Burgyan J. 2010. Polerovirus protein P0 prevents the assembly of small RNA-containing RISC complexes and leads to degradation of ARGONAUTE1. Plant J 62: 463–472.

26. VarallyayE, ValocziA, AgyiA, BurgyanJ, HaveldaZ (2010) Plant virus-mediated induction of miR168 is associated with repression of ARGONAUTE1 accumulation. EMBO J 29: 3507–3519.

27. AnandalakshmiR, MaratheR, GeX, HerrJMJr, MauC, et al. (2000) A calmodulin-related protein that suppresses posttranscriptional gene silencing in plants. Science 290: 142–144.

28. EndresMW, GregoryBD, GaoZ, ForemanAW, MlotshwaS, et al. (2010) Two plant viral suppressors of silencing require the ethylene-inducible host transcription factor RAV2 to block RNA silencing. PloS Pathog 6: e1000729 doi: 10.1371/journal.ppat.1000729

29. HoeflichKP, IkuraM (2002) Calmodulin in action: diversity in target recognition and activation mechanisms. Cell 108: 739–742.

30. KimMC, ChungWS, YunDJ, ChoMJ (2009) Calcium and calmodulin-mediated regulation of gene expression in plants. Mol Plant 2: 13–21.

31. ChenX (2009) Small RNAs and their roles in plant development. Annu Rev Cell Dev Biol 25: 21–44.

32. JeskeH (2009) Geminiviruses. Curr Top Microbiol Immunol 331: 185–226.

33. BisaroDM (2006) Silencing suppression by geminivirus proteins. Virology 344: 158–168.

34. HohnT, VazquezF (2011) RNA silencing pathways of plants: silencing and its suppression by plant DNA viruses. Biochim Biophys Acta 1809: 588–600.

35. AkbergenovR, Si-AmmourA, BlevinsT, AminI, KutterC, et al. (2006) Molecular characterization of geminivirus-derived small RNAs in different plant species. Nucleic Acids Res 34: 462–471.

36. MuangsanN, BéclinC, VaucheretH, RobertsonD (2004) Geminivirus VIGS of endogenous genes requires SGS2/SDE1 and SGS3 and defines a new branch in the genetic pathway for silencing in plants. Plant J 38: 1004–1014.

37. BriddonRW, BullSE, AminI, MansoorS, BedfordID, RishiN, et al. (2003) Diversity of DNA β, a satellite molecule associated with some monopartite begomoviruses. Virology 312: 106–121.

38. ZhouX, XieY, TaoX, ZhangZ, LiZ, et al. (2003) Characterization of DNA β associated with begomoviruses in China and evidence for co-evolution with their cognate viral DNA-A. J Gen Virol 84: 237–47.

39. CuiX, TaoX, XieY, FauquetCM, ZhouX (2004) A DNA β associated with Tomato yellow leaf curl China virus is required for symptom induction. J Virol 78: 13966–13974.

40. DingC, QingL, LiZ, LiuY, QianY, ZhouX (2009) Genetic determinants of symptoms on viral DNA satellites. Appl Environ Microbiol 2009 75: 5380–5389.

41. QaziJ, AminI, MansoorS, IqbalMJ, BriddonRW (2007) Contribution of the satellite encoded gene βC1 to cotton leaf curl disease symptoms. Virus Res 128: 135–139.

42. SaundersK, NormanA, GucciardoS, StanleyJ (2004) The DNA β satellite component associated with ageratum yellow vein disease encodes an essential pathogenicity protein (βC1). Virology 324: 37–47.

43. AminI, HussainK, AkbergenovR, YadavJS, QaziJ, et al. (2011) Suppressors of RNA silencing encoded by the components of the cotton leaf curl begomovirus-betasatellite complex. Mol Plant Microbe Interact 24: 973–983.

44. CuiX, LiG, WangD, HuD, ZhouX (2005) A begomovirus DNA β-encoded protein binds DNA, functions as a suppressor of RNA silencing, and targets the cell nucleus. J Virol 79: 10764–10775.

45. EiniO, DograSC, DryIB, RandlesJW (2012) Silencing suppressor activity of a begomovirus DNA β encoded protein and its effect on heterologous helper virus replication. Virus Res 167: 97–101.

46. GopalP, Pravin KumarP, SinilalB, JoseJ, Kasin YadunandamA, et al. (2007) Differential roles of C4 and βC1 in mediating suppression of post-transcriptional gene silencing: Evidence for transactivation by the C2 of Bhendi yellow vein mosaic virus, a monopartite begomovirus. Virus Res 123: 9–18.

47. KonT, SharmaP, IkegamiM (2007) Suppressor of RNA silencing encoded by the monopartite tomato leaf curl Java begomovirus. Arch Virol 152: 1273–1282.

48. SharmaP, IkegamiM, KonT (2010) Identification of the virulence factors and suppressors of posttranscriptional gene silencing encoded by Ageratum yellow vein virus, a monopartite begomovirus. Virus Res 149: 19–27.

49. ShuklaR, DalalS, MalathiVG (2013) Suppressors of RNA silencing encoded by tomato leaf curl betasatellites. J Biosci 38: 45–51.

50. BrignetiG, VoinnetO, LiWX, JiLH, DingSW, et al. (1998) Viral pathogenicity determinants are suppressors of transgene silencing in Nicotiana benthamiana. EMBO J 17: 6739–6746.

51. JohansenLK, CarringtonJC (2001) Silencing on the spot. Induction and suppression of RNA silencing in the Agrobacterium-mediated transient expression system. Plant Physiol 126: 930–938.

52. HuangC, XieY, ZhouX (2009) Efficient virus-induced gene silencing in plants using a modified geminivirus DNA1 component. Plant Biotechnol J 7: 254–265.

53. HuangCJ, ZhangT, LiFF, ZhangXY, ZhouXP (2011) Development and application of an efficient virus-induced gene silencing system in Nicotiana tabacum using geminivirus alphasatellite. J Zhejiang Univ Sci B 12: 83–92.

54. Dinesh-KumarSP, AnandalakshmiR, MaratheR, SchiffM, LiuY (2003) Virus-induced gene silencing. Methods Mol Biol 236: 287–94.

55. YangJY, IwasakiM, MachidaC, MachidaY, ZhouX, et al. (2008) βC1, the pathogenicity factor of TYLCCNV, interacts with AS1 to alter leaf development and suppress selective jasmonic acid responses. Genes Dev 22: 2564–2577.

56. GlazovE, PhillipsK, BudziszewskiGJ, MeinsF, LevinJZ (2003) A gene encoding an RNase D exonuclease-like protein is required for post-transcriptional silencing in Arabidopsis. Plant J 35: 342–349.

57. GyI, GasciolliV, LauresserguesD, MorelJB, GombertJ, ProuxF, et al. (2007) Arabidopsis FIERY1, XRN2, and XRN3 are endogenous RNA silencing suppressors. Plant Cell 19: 3451–3461.

58. Le MassonI, JauvionV, BouteillerN, RivardM, ElmayanT, VaucheretH (2012) Mutations in the Arabidopsis H3K4me2/3 demethylase JMJ14 suppress posttranscriptional gene silencing by decreasing transgene transcription. Plant Cell 24: 3603–3612.

59. LiX, QianW, ZhaoY, WangC, ShenJ, et al. (2012) Antisilencing role of the RNA-directed DNA methylation pathway and a histone acetyltransferase in Arabidopsis. Proc Natl Acad Sci U S A 109: 11425–11430.

60. MorenoAB, Martínez de AlbaAE, BardouF, CrespiMD, VaucheretH, et al. (2013) Cytoplasmic and nuclear quality control and turnover of single-stranded RNA modulate post-transcriptional gene silencing in plants. Nucleic Acids Res 41: 4699–4708.

61. NakaharaKS, MasutaC, YamadaS, ShimuraH, KashiharaY, et al. (2012) Tobacco calmodulin-like protein provides secondary defense by binding to and directing degradation of virus RNA silencing suppressors. Proc Natl Acad Sci U S A 109: 10113–10118.

62. Roth BM. 2006. Viral and endogenous suppressors of RNA silencing in Arabidopsis small RNA pathways. University of South Carolina. Ph.D thesis. p73–74.

63. MlotshwaS, VerverJ, Sithole-NiangI, GopinathK, CaretteJ, van KammenA, et al. (2002) Subcellular location of the helper component-proteinase of Cowpea aphid-borne mosaic virus. Virus Genes 25: 207–216.

64. Cheng X (2010) Biochemical analysis of βC1 encoded by betasatellite of begomoviruses. Zhejiang University. Ph.D thesis. p72.

65. TalianskyM, KimSH, MayoMA, KalininaNO, FraserG, et al. (2004) Escape of a plant virus from amplicon-mediated RNA silencing is associated with biotic or abiotic stress. Plant J 39: 194–205.

66. YangX, XieY, RajaP, LiS, WolfJN, et al. (2011) Suppression of methylation-mediated transcriptional gene silencing by βC1-SAHH protein interaction during geminivirus-betasatellite infection. PLoS Pathog 7: e1002329 doi: 10.1371/journal.ppat.1002329

67. WuQ, WangX, DingSW (2010) Viral suppressors of RNA-based viral immunity: host targets. Cell Host Microbe 8: 12–15.

68. LiY, LuJ, HanY, FanX, DingSW (2013) RNA interference functions as an antiviral immunity mechanism in mammals. Science 342: 231–234.

69. MaillardPV, CiaudoC, MarchaisA, LiY, JayF, DingSW, VoinnetO (2013) Antiviral RNA interference in mammalian cells. Science 342: 235–238.

70. JaubertM, BhattacharjeeS, MelloAF, PerryKL, MoffettP (2011) ARGONAUTE2 mediates RNA-silencing antiviral defenses against Potato virus X in Arabidopsis. Plant Physiol 156: 1556–1564.

71. FukunagaR, DoudnaJA (2009) dsRNA with 5′ overhangs contributes to endogenous and antiviral RNA silencing pathways in plants. EMBO J 28: 545–555.

72. GlickE, ZrachyaA, LevyY, MettA, GidoniD, et al. (2008) Interaction with host SGS3 is required for suppression of RNA silencing by tomato yellow leaf curl virus V2 protein. Proc Natl Acad Sci U S A 105: 157–161.

73. NaimF, NakasugiK, CrowhurstRN, HilarioE, ZwartAB, et al. (2012) Advanced engineering of lipid metabolism in Nicotiana benthamiana using a draft genome and the V2 viral silencing-suppressor protein. PLoS One 7: e52717 doi: 10.1371/journal.pone.0052717

74. AreggerM, BorahBK, SeguinJ, RajeswaranR, GubaevaEG, et al. (2012) Primary and secondary siRNAs in geminivirus-induced gene silencing. PLoS Pathog 8 (9) e1002941 doi: 10.1371/journal.ppat.1002941

75. VerlaanMG, HuttonSF, IbrahemRM, KormelinkR, VisserRG, et al. (2013) The tomato yellow leaf curl virus resistance genes Ty-1 and Ty-3 are allelic and code for DFDGD-class RNA dependent RNA polymerases. PLoS Genet 9: e1003399 doi: 10.1371/journal.pgen.1003399

76. LuoZ, ChenZ (2007) Improperly terminated, unpolyadenylated mRNA of sense transgenes is targeted by RDR6-mediated RNA silencing in Arabidopsis. Plant Cell 19: 943–958.

77. RajaP, SanvilleBC, BuchmannRC, BisaroDM (2008) Viral genome methylation as an epigenetic defense against geminiviruses. J Virol 82: 8997–9007.

78. GuoM, ThomasJ, CollinsG, TimmermansMC (2008) Direct repression of KNOX loci by the ASYMMETRIC LEAVES1 complex of Arabidopsis. Plant Cell 20: 48–58.

79. LiH, XuL, WangH, YuanZ, CaoX, et al. (2005) The putative RNA-dependent RNA polymerase RDR6 acts synergistically with ASYMMETRIC LEAVES1 and 2 to repress BREVIPEDICELLUS and MicroRNA165/166 in Arabidopsis leaf development. Plant Cell 17: 2157–2171.

80. XuL, YangL, PiL, LiuQ, LingQ, et al. (2006) Genetic interaction between the AS1-AS2 and RDR6-SGS3-AGO7 pathways for leaf morphogenesis. Plant Cell Physiol 47: 853–863.

81. HanHJ, ParkHC, ByunHJ, LeeSM, KimHS, et al. (2012) The transcriptional repressor activity of ASYMMETRIC LEAVES1 is inhibited by direct interaction with calmodulin in Arabidopsis. Plant Cell Environ 35: 1969–1982.

82. XiongR, WuJ, ZhouY, ZhouX (2009) Characterization and subcellular localization of an RNA silencing suppressor encoded by Rice stripe tenuivirus. Virology 387: 29–40.