PIWI Associated siRNAs and piRNAs Specifically Require the HEN1 Ortholog

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Small RNAs—including piRNAs, miRNAs, and endogenous siRNAs—bind Argonaute proteins to form RNA silencing complexes that target coding genes, transposons, and aberrant RNAs. To assess the requirements for endogenous siRNA formation and activity in Caenorhabditis elegans, we developed a GFP-based sensor for the endogenous siRNA 22G siR-1, one of a set of abundant siRNAs processed from a precursor RNA mapping to the X chromosome, the X-cluster. Silencing of the sensor is also dependent on the partially complementary, unlinked 26G siR-O7 siRNA. We show that 26G siR-O7 acts in trans to initiate 22G siRNA formation from the X-cluster. The presence of several mispairs between 26G siR-O7 and the X-cluster mRNA, as well as mutagenesis of the siRNA sensor, indicates that siRNA target recognition is permissive to a degree of mispairing. From a candidate reverse genetic screen, we identified several factors required for 22G siR-1 activity, including the chromatin factors mes-4 and gfl-1, the Argonaute ergo-1, and the 3′ methyltransferase henn-1. Quantitative RT–PCR of small RNAs in a henn-1 mutant and deep sequencing of methylated small RNAs indicate that siRNAs and piRNAs that associate with PIWI clade Argonautes are methylated by HENN-1, while siRNAs and miRNAs that associate with non-PIWI clade Argonautes are not. Thus, PIWI-class Argonaute proteins are specifically adapted to associate with methylated small RNAs in C. elegans.

Vyšlo v časopise: PIWI Associated siRNAs and piRNAs Specifically Require the HEN1 Ortholog. PLoS Genet 8(4): e32767. doi:10.1371/journal.pgen.1002616
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.pgen.1002616

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Zdroje

1. CzechBHannonGJ 2011 Small RNA sorting: matchmaking for Argonautes. Nat Rev Genet 12 19 31

2. ToliaNHJoshua-TorL 2007 Slicer and the argonautes. Nat Chem Biol 3 36 43

3. GrishokAPasquinelliAEConteDLiNParrishS 2001 Genes and mechanisms related to RNA interference regulate expression of the small temporal RNAs that control C. elegans developmental timing. Cell 106 23 34

4. BatistaPJRubyJGClaycombJMChiangRFahlgrenN 2008 PRG-1 and 21U-RNAs interact to form the piRNA complex required for fertility in C. elegans. Mol Cell 31 67 78

5. DasPPBagijnMPGoldsteinLDWoolfordJRLehrbachNJ 2008 Piwi and piRNAs act upstream of an endogenous siRNA pathway to suppress Tc3 transposon mobility in the Caenorhabditis elegans germline. Mol Cell 31 79 90

6. YigitEBatistaPJBeiYPangKMChenCC 2006 Analysis of the C. elegans Argonaute family reveals that distinct Argonautes act sequentially during RNAi. Cell 127 747 757

7. GuWShirayamaMConteDJrVasaleJBatistaPJ 2009 Distinct argonaute-mediated 22G-RNA pathways direct genome surveillance in the C. elegans germline. Mol Cell 36 231 244

8. ConineCCBatistaPJGuWClaycombJMChavesDA 2010 Argonautes ALG-3 and ALG-4 are required for spermatogenesis-specific 26G-RNAs and thermotolerant sperm in Caenorhabditis elegans. Proc Natl Acad Sci U S A 107 3588 3593

9. HanTManoharanAPHarkinsTTBouffardPFitzpatrickC 2009 26G endo-siRNAs regulate spermatogenic and zygotic gene expression in Caenorhabditis elegans. Proc Natl Acad Sci U S A 106 18674 18679

10. VasaleJJGuWThiviergeCBatistaPJClaycombJM 2010 Sequential rounds of RNA-dependent RNA transcription drive endogenous small-RNA biogenesis in the ERGO-1/Argonaute pathway. Proc Natl Acad Sci U S A 107 3582 3587

11. RubyJGJanCPlayerCAxtellMJLeeW 2006 Large-scale sequencing reveals 21U-RNAs and additional microRNAs and endogenous siRNAs in C. elegans. Cell 127 1193 1207

12. WangGReinkeV 2008 A C. elegans Piwi, PRG-1, regulates 21U-RNAs during spermatogenesis. Curr Biol 18 861 867

13. ZhangCMontgomeryTAGabelHWFischerSEPhillipsCM 2011 mut-16 and other mutator class genes modulate 22G and 26G siRNA pathways in Caenorhabditis elegans. Proc Natl Acad Sci U S A 108 1201 1208

14. KettingRF 2011 The many faces of RNAi. Dev Cell 20 148 161

15. BernsteinECaudyAAHammondSMHannonGJ 2001 Role for a bidentate ribonuclease in the initiation step of RNA interference. Nature 409 363 366

16. KettingRFFischerSEBernsteinESijenTHannonGJ 2001 Dicer functions in RNA interference and in synthesis of small RNA involved in developmental timing in C. elegans. Genes Dev 15 2654 2659

17. HammondSMBernsteinEBeachDHannonGJ 2000 An RNA-directed nuclease mediates post-transcriptional gene silencing in Drosophila cells. Nature 404 293 296

18. GentJILammATPavelecDMManiarJMParameswaranP 2010 Distinct phases of siRNA synthesis in an endogenous RNAi pathway in C. elegans soma. Mol Cell 37 679 689

19. ClaycombJMBatistaPJPangKMGuWVasaleJJ 2009 The Argonaute CSR-1 and its 22G-RNA cofactors are required for holocentric chromosome segregation. Cell 139 123 134

20. van WolfswinkelJCClaycombJMBatistaPJMelloCCBerezikovE 2009 CDE-1 affects chromosome segregation through uridylation of CSR-1-bound siRNAs. Cell 139 135 148

21. GuangSBochnerAFPavelecDMBurkhartKBHardingS 2008 An Argonaute transports siRNAs from the cytoplasm to the nucleus. Science 321 537 541

22. GuangSBochnerAFBurkhartKBBurtonNPavelecDM 2010 Small regulatory RNAs inhibit RNA polymerase II during the elongation phase of transcription. Nature 465 1097 1101

23. BilliACAlessiAFKhivansaraVHanTFreebergM 2012 The Caenorhabditis elegans HEN1 ortholog, HENN-1, methylates and stabilizes select subclasses of germline small RNAs. PLoS Genet 8 e1002617 doi:10.1371/journal.pgen.1002617

24. KammingaLMvan WolfswinkelJCLuteijnMJKaaijLJTBagijnMP 2012 Differential impact of the Hen1 homolog HENN-1 on 21U and 26G RNAs in the germline of Caenorhabditis elegans. PLoS Genet In press

25. Frokjaer-JensenCDavisMWHopkinsCENewmanBJThummelJM 2008 Single-copy insertion of transgenes in Caenorhabditis elegans. Nat Genet 40 1375 1383

26. AmbrosVLeeRCLavanwayAWilliamsPTJewellD 2003 MicroRNAs and other tiny endogenous RNAs in C. elegans. Curr Biol 13 807 818

27. FischerSEButlerMDPanQRuvkunG 2008 Trans-splicing in C. elegans generates the negative RNAi regulator ERI-6/7. Nature 455 491 496

28. KimJKGabelHWKamathRSTewariMPasquinelliA 2005 Functional genomic analysis of RNA interference in C. elegans. Science 308 1164 1167

29. TabaraHSarkissianMKellyWGFleenorJGrishokA 1999 The rde-1 gene, RNA interference, and transposon silencing in C. elegans. Cell 99 123 132

30. SimmerFTijstermanMParrishSKoushikaSPNonetML 2002 Loss of the putative RNA-directed RNA polymerase RRF-3 makes C. elegans hypersensitive to RNAi. Curr Biol 12 1317 1319

31. SijenTFleenorJSimmerFThijssenKLParrishS 2001 On the role of RNA amplification in dsRNA-triggered gene silencing. Cell 107 465 476

32. SijenTSteinerFAThijssenKLPlasterkRH 2007 Secondary siRNAs result from unprimed RNA synthesis and form a distinct class. Science 315 244 247

33. PakJFireA 2007 Distinct populations of primary and secondary effectors during RNAi in C. elegans. Science 315 241 244

34. BartelDP 2009 MicroRNAs: target recognition and regulatory functions. Cell 136 215 233

35. WangYJuranekSLiHShengGTuschlT 2008 Structure of an argonaute silencing complex with a seed-containing guide DNA and target RNA duplex. Nature 456 921 926

36. GersteinMBLuZJVan NostrandELChengCArshinoffBI 2010 Integrative analysis of the Caenorhabditis elegans genome by the modENCODE project. Science 330 1775 1787

37. FischerSEMontgomeryTAZhangCFahlgrenNBreenPC 2011 The ERI-6/7 Helicase Acts at the First Stage of an siRNA Amplification Pathway That Targets Recent Gene Duplications. PLoS Genet 7 e1002369 doi:10.1371/journal.pgen.1002369

38. Frokjaer-JensenCDavisMWHollopeterGTaylorJHarrisTW 2010 Targeted gene deletions in C. elegans using transposon excision. Nature methods 7 451 453

39. YuBYangZLiJMinakhinaSYangM 2005 Methylation as a crucial step in plant microRNA biogenesis. Science 307 932 935

40. KirinoYMourelatosZ 2007 Mouse Piwi-interacting RNAs are 2′-O-methylated at their 3′ termini. Nature structural & molecular biology 14 347 348

41. OharaTSakaguchiYSuzukiTUedaHMiyauchiK 2007 The 3′ termini of mouse Piwi-interacting RNAs are 2′-O-methylated. Nature structural & molecular biology 14 349 350

42. HorwichMDLiCMatrangaCVaginVFarleyG 2007 The Drosophila RNA methyltransferase, DmHen1, modifies germline piRNAs and single-stranded siRNAs in RISC. Curr Biol 17 1265 1272

43. SaitoKSakaguchiYSuzukiTSiomiHSiomiMC 2007 Pimet, the Drosophila homolog of HEN1, mediates 2′-O-methylation of Piwi- interacting RNAs at their 3′ ends. Genes & development 21 1603 1608

44. KurthHMMochizukiK 2009 2′-O-methylation stabilizes Piwi-associated small RNAs and ensures DNA elimination in Tetrahymena. RNA 15 675 685

45. CuppenEGortEHazendonkEMuddeJvan de BeltJ 2007 Efficient target-selected mutagenesis in Caenorhabditis elegans: toward a knockout for every gene. Genome Res 17 649 658

46. LiJYangZYuBLiuJChenX 2005 Methylation protects miRNAs and siRNAs from a 3′-end uridylation activity in Arabidopsis. Curr Biol 15 1501 1507

47. YuBChenX 2010 Analysis of miRNA Modifications. Methods Mol Biol 592 137 148

48. GhildiyalMSeitzHHorwichMDLiCDuT 2008 Endogenous siRNAs derived from transposons and mRNAs in Drosophila somatic cells. Science 320 1077 1081

49. AmeresSLHorwichMDHungJHXuJGhildiyalM 2010 Target RNA-directed trimming and tailing of small silencing RNAs. Science 328 1534 1539

50. AllenEXieZGustafsonAMCarringtonJC 2005 microRNA-directed phasing during trans-acting siRNA biogenesis in plants. Cell 121 207 221

51. AxtellMJJanCRajagopalanRBartelDP 2006 A two-hit trigger for siRNA biogenesis in plants. Cell 127 565 577

52. CorreaRLSteinerFABerezikovEKettingRF 2010 MicroRNA-directed siRNA biogenesis in Caenorhabditis elegans. PLoS Genet 6 e1000903 doi:10.1371/journal.pgen.1000903

53. HowellMDFahlgrenNChapmanEJCumbieJSSullivanCM 2007 Genome-wide analysis of the RNA-DEPENDENT RNA POLYMERASE6/DICER-LIKE4 pathway in Arabidopsis reveals dependency on miRNA- and tasiRNA-directed targeting. The Plant cell 19 926 942

54. SongJJLiuJToliaNHSchneidermanJSmithSK 2003 The crystal structure of the Argonaute2 PAZ domain reveals an RNA binding motif in RNAi effector complexes. Nat Struct Biol 10 1026 1032

55. LingelASimonBIzaurraldeESattlerM 2004 Nucleic acid 3′-end recognition by the Argonaute2 PAZ domain. Nat Struct Mol Biol 11 576 577

56. MaJBYeKPatelDJ 2004 Structural basis for overhang-specific small interfering RNA recognition by the PAZ domain. Nature 429 318 322

57. TianYSimanshuDKMaJBPatelDJ 2011 Structural basis for piRNA 2′-O-methylated 3′-end recognition by Piwi PAZ (Piwi/Argonaute/Zwille) domains. Proc Natl Acad Sci U S A 108 903 910

58. PhillipsCMMcDonaldKLDernburgAF 2009 Cytological analysis of meiosis in Caenorhabditis elegans. Methods Mol Biol 558 171 195

59. FahlgrenNSullivanCMKasschauKDChapmanEJCumbieJS 2009 Computational and analytical framework for small RNA profiling by high-throughput sequencing. RNA 15 992 1002

60. Griffiths-JonesS 2010 miRBase: microRNA sequences and annotation. Current protocols in bioinformatics/editoral board, Andreas D Baxevanis [et al] Chapter 12 Unit 12 19 11-10

61. KamathRSFraserAGDongYPoulinGDurbinR 2003 Systematic functional analysis of the Caenorhabditis elegans genome using RNAi. Nature 421 231 237

62. RualJFCeronJKorethJHaoTNicotAS 2004 Toward improving Caenorhabditis elegans phenome mapping with an ORFeome-based RNAi library. Genome Res 14 2162 2168

63. LlaveCXieZKasschauKDCarringtonJC 2002 Cleavage of Scarecrow-like mRNA targets directed by a class of Arabidopsis miRNA. Science 297 2053 2056

64. LarkinMABlackshieldsGBrownNPChennaRMcGettiganPA 2007 Clustal W and Clustal X version 2.0. Bioinformatics 23 2947 2948