Application of Systemic Transcriptional Gene Silencing for Plant Breeding

  • Songling Bai
  • Takeo Harada
  • Atsushi Kasai


Small interfering RNA (siRNA)-mediated gene silencing has been observed in eukaryotes across all kingdoms from fungi to mammals. In plants, this phenomenon influences resistance to pathogenic viruses, suppression of transgene expression, and the inactivation of transposable elements. Recent studies have revealed that double-stranded RNA-derived siRNAs are able to induce systemic transcriptional gene silencing (TGS) in graft partners. In particular, when the scion is used as the siRNA donor, the roots exhibit strong systemic TGS, especially the lateral roots. Such gene silencing can be maintained through in vitro regeneration and is heritable. We developed a novel method for transforming plants using this process. The expression of the target gene can be arrested without inserting exogenous DNA into the genome of the target organism. We herein review the recent advances in research related to systemic TGS. We also describe the potential utility of systemic TGS for plant breeding.


Transcriptional gene silencing RNA-directed DNA methylation Epigenetics Grafting Genetically modified organism Plant breeding technique 


  1. Allison R, Thompson C, Ahlquist P (1990) Regeneration of a functional RNA virus genome by recombination between deletion mutants and requirement for cowpea chlorotic mottle virus 3a and coat genes for systemic infection. Proc Natl Acad Sci 87:1820–1824CrossRefGoogle Scholar
  2. Aloni B, Karni L, Deventurero G, Levin Z, Cohen R, Katzir N, Lotan-Pompan M, Edelstein M, Aktas H, Turhan E, Joel DM, Horev C, Kapulnik Y (2008) Physiological and biochemical changes at the rootstock-scion interface in graft combinations between Cucurbita rootstocks and a melon scion. J Hortic Sci Biotechnol 83:777–783CrossRefGoogle Scholar
  3. Axtell M, Merchant S (2013) Classification and comparison of small RNAs from plants. Annu Rev Plant Biol 64:137–159CrossRefGoogle Scholar
  4. Bai S, Kasai A, Yamada K, Li T, Harada T (2011) A mobile signal transported over a long distance induces systemic transcriptional gene silencing in a grafted partner. J Exp Bot 62:4561–4570CrossRefGoogle Scholar
  5. Bartel DP (2004) MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 116:281–297CrossRefGoogle Scholar
  6. Bartel DP (2009) MicroRNAs: target recognition and regulatory functions. Cell 136:215–233CrossRefGoogle Scholar
  7. Baulcombe D (2004) RNA silencing in plants. Nature 431:356–363CrossRefGoogle Scholar
  8. Becker C, Hagmann J, Muller J, Koenig D, Stegle O, Borgwardt K, Weigel D (2011) Spontaneous epigenetic variation in the Arabidopsis thaliana methylome. Nature 480:245–249CrossRefGoogle Scholar
  9. Blevins T, Pontvianne F, Cocklin R, Podicheti R, Chandrasekhara C, Yerneni S, Braun C, Lee B, Rusch D, Mockaitis K, Tang H, Pikaard CS (2014) A two-step process for epigenetic inheritance in Arabidopsis. Mol Cell 54:30–42CrossRefGoogle Scholar
  10. Bond DM, Baulcombe DC (2015) Epigenetic transitions leading to heritable, RNA-mediated de novo silencing in Arabidopsis thaliana. Proc Natl Acad Sci U S A 112:917–922CrossRefGoogle Scholar
  11. Brosnan CA, Mitter N, Christie M, Smith NA, Waterhouse PM, Carroll BJ (2007) Nuclear gene silencing directs reception of long-distance mRNA silencing in Arabidopsis. Proc Natl Acad Sci U S A 104:14741–14746CrossRefGoogle Scholar
  12. Chen ZJ (2013) Genomic and epigenetic insights into the molecular bases of heterosis. Nat Rev Genet 14:471–482CrossRefGoogle Scholar
  13. Covey SN, Al-Kaff NS, Lángara A, Turner DS (1997) Plants combat infection by gene silencing. Nature 385:781–782CrossRefGoogle Scholar
  14. Cutting JGM, Lyne MC (1993) Girdling and the reduction in Shoot Xylem Sap concentrations of Cytokinins and Gibberellins in Peach. J Hortic Sci 68:619–626CrossRefGoogle Scholar
  15. Daxinger L, Kanno T, Bucher E, van der Winden J, Naumann U, Matzke AJM, Matzke M (2009) A stepwise pathway for biogenesis of 24-nt secondary siRNAs and spreading of DNA methylation. EMBO J 28:48–57CrossRefGoogle Scholar
  16. De Smet I (2011) Lateral root initiation: one step at a time. New Phytol 193:867–873CrossRefGoogle Scholar
  17. Fei Q, Xia R, Meyers B (2013) Phased, secondary, small interfering RNAs in posttranscriptional regulatory networks. Plant Cell 25:2400–2415CrossRefGoogle Scholar
  18. Fire A, Xu SQ, Montgomery MK, Kostas SA, Driver SE, Mello CC (1998) Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391:806–811CrossRefGoogle Scholar
  19. Gohlke J, Mosher RA (2015) Exploiting mobile RNA silencing for crop improvement. Am J Bot 102:1399–1400CrossRefGoogle Scholar
  20. Goldschmidt EE (2014) Plant grafting: new mechanisms, evolutionary implications. Front Plant Sci 5:727CrossRefGoogle Scholar
  21. Guo S, Kemphues KJ (1995) Par-1, a gene required for establishing polarity in C. elegans embryos, encodes a putative Ser/Thr kinase that is asymmetrically distributed. Cell 81:611–620CrossRefGoogle Scholar
  22. Han X, Kumar D, Chen H, Wu S, Kim JY (2014) Transcription factor-mediated cell-to-cell signalling in plants. J Exp Bot 65:1737–1749CrossRefGoogle Scholar
  23. Hannapel DJ, Sharma P, Lin T (2013) Phloem-mobile messenger RNAs and root development. Front Plant Sci 4:257PubMedPubMedCentralGoogle Scholar
  24. Harada T (2010) Grafting and RNA transport via phloem tissue in horticultural plants. Sci Hortic 125:545–550CrossRefGoogle Scholar
  25. Haroldsen VM, Chi-Ham CL, Bennett AB (2012) Transgene mobilization and regulatory uncertainty for non-GE fruit products of transgenic rootstocks. J Biotechnol 161:349–353CrossRefGoogle Scholar
  26. Heap B (2013) Europe should rethink its stance on GM crops. Nature 498:409–409CrossRefGoogle Scholar
  27. Heilersig BHJB, Loonen AEHM, Janssen EM, Wolters A-MA, Visser RGF (2006) Efficiency of transcriptional gene silencing of GBSSI in potato depends on the promoter region that is used in an inverted repeat. Mol Gen Genomics 275:437–449CrossRefGoogle Scholar
  28. Himber C, Dunoyer P, Moissiard G, Ritzenthaler C, Voinnet O (2003) Transitivity-dependent and -independent cell-to-cell movement of RNA silencing. EMBO J 22:4523–4533CrossRefGoogle Scholar
  29. Holoch D, Moazed D (2015) RNA-mediated epigenetic regulation of gene expression. Nat Rev Genet 16:71–84CrossRefGoogle Scholar
  30. Jones L, Hamilton AJ, Voinnet O, Thomas CL, Maule AJ, Baulcombe DC (1999) RNA-DNA interactions and DNA methylation in post-transcriptional gene silencing. Plant Cell 11:2291–2301CrossRefGoogle Scholar
  31. Jones L, Ratcliff F, Baulcombe DC (2001) RNA-directed transcriptional gene silencing in plants can be inherited independently of the RNA trigger and requires Met1 for maintenance. Curr Biol 11:747–757CrossRefGoogle Scholar
  32. Kalantidis K, Schumacher HT, Alexiadis T, Helm JM (2008) RNA silencing movement in plants. Biol Cell 100:13–26CrossRefGoogle Scholar
  33. Kanazawa A, Inaba J, Shimura H, Otagaki S, Tsukahara S, Matsuzawa A, Kim BM, Goto K, Masuta C (2011) Virus-mediated efficient induction of epigenetic modifications of endogenous genes with phenotypic changes in plants. Plant J 65:156–168CrossRefGoogle Scholar
  34. Kasai A, Bai S, Hojo H, Harada T (2016) Epigenome editing of potato by grafting using transgenic tobacco as siRNA donor. PLoS One 11:e0161729CrossRefGoogle Scholar
  35. Law JA, Jacobsen SE (2010) Establishing, maintaining and modifying DNA methylation patterns in plants and animals. Nat Rev Genet 11:204–220CrossRefGoogle Scholar
  36. Lewsey MG, Hardcastle TJ, Melnyk CW, Molnar A, Valli A, Urich MA, Nery JR, Baulcombe DC, Ecker JR (2016) Mobile small RNAs regulate genome-wide DNA methylation. Proc Natl Acad Sci U S A 113:E801–E810CrossRefGoogle Scholar
  37. Liang DC, White RG, Waterhouse PM (2012) Gene silencing in Arabidopsis spreads from the root to the shoot, through a gating barrier, by template-dependent, nonvascular, cell-to-cell movement. Plant Physiol 159:984–1000CrossRefGoogle Scholar
  38. Lloret P, Casero P (2002) Lateral root initiation. In: Plant roots. CRC Press, Boca Raton, pp 127–155. CrossRefGoogle Scholar
  39. Lough TJ, Lucas WJ (2006) Integrative plant biology: role of phloem long-distance macromolecular trafficking. Annu Rev Plant Biol 57:203–232CrossRefGoogle Scholar
  40. Lu R, Martin-Hernandez AM, Peart JR, Malcuit I, Baulcombe DC (2003) Virus-induced gene silencing in plants. Methods 30:296–303CrossRefGoogle Scholar
  41. Lusser M, Parisi C, Plan D, Rodríguez-Cerezo E (2012) Deployment of new biotechnologies in plant breeding. Nat Biotechnol 30:231–239CrossRefGoogle Scholar
  42. Mark Cigan A, Unger-Wallace E, Haug-Collet K (2005) Transcriptional gene silencing as a tool for uncovering gene function in maize. Plant J 43:929–940CrossRefGoogle Scholar
  43. Martin A, Troadec C, Boualem A, Rajab M, Fernandez R, Morin H, Pitrat M, Dogimont C, Bendahmane A (2009) A transposon-induced epigenetic change leads to sex determination in melon. Nature 461:1135–1138CrossRefGoogle Scholar
  44. Matsuda Y, Liang G, Zhu Y, Ma F, Nelson RS, Ding B (2002) The Commelina yellow mottle virus promoter drives companion-cell-specific gene expression in multiple organs of transgenic tobacco. Protoplasma 220:51–58CrossRefGoogle Scholar
  45. Melnyk CW, Molnar A, Bassett A, Baulcombe DC (2011a) Mobile 24 nt small RNAs direct transcriptional gene silencing in the root meristems of Arabidopsis thaliana. Curr Biol 21:1678–1683CrossRefGoogle Scholar
  46. Melnyk CW, Molnar A, Baulcombe DC (2011b) Intercellular and systemic movement of RNA silencing signals. EMBO J 30:3553–3563CrossRefGoogle Scholar
  47. Mermigka G, Verret F, Kalantidis K (2016) RNA silencing movement in plants. J Integr Plant Biol 58:328–342CrossRefGoogle Scholar
  48. Miguel C, Marum L (2011) An epigenetic view of plant cells cultured in vitro: somaclonal variation and beyond. J Exp Bot 62:3713–3725CrossRefGoogle Scholar
  49. Mlotshwa S, Voinnet O, Mette MF, Matzke M, Vaucheret H, Ding SW, Pruss G, Vance VB (2002) RNA silencing and the mobile silencing signal. Plant Cell 14:S289–S301CrossRefGoogle Scholar
  50. Molnar A, Csorba T, Lakatos L, Varallyay E, Lacomme C, Burgyan J (2005) Plant virus-derived small interfering RNAs originate predominantly from highly structured single-stranded viral RNAs. J Virol 79:7812–7818CrossRefGoogle Scholar
  51. Molnar A, Schwach F, Studholme DJ, Thuenemann EC, Baulcombe DC (2007) miRNAs control gene expression in the single-cell alga Chlamydomonas reinhardtii. Nature 447:1126–U1115CrossRefGoogle Scholar
  52. Molnar A, Melnyk CW, Bassett A, Hardcastle TJ, Dunn R, Baulcombe DC (2010) Small silencing RNAs in plants are mobile and direct epigenetic modification in recipient cells. Science 328:872–875CrossRefGoogle Scholar
  53. Mourrain P, van Blokland R, Kooter JM, Vaucheret H (2007) A single transgene locus triggers both transcriptional and post-transcriptional silencing through double-stranded RNA production. Planta 225:365–379CrossRefGoogle Scholar
  54. Mudge K, Janick J, Scofield S, Goldschmidt EE (2009) A history of grafting. Hortic Rev 35:437–493. CrossRefGoogle Scholar
  55. Napoli C, Lemieux C, Jorgensen R (1990) Introduction of a chimeric chalcone synthase gene into petunia results in reversible co-suppression of homologous genes in trans. Plant Cell 2:279–289CrossRefGoogle Scholar
  56. Pina A, Errea P (2005) A review of new advances in mechanism of graft compatibility-incompatibility. Sci Hortic 106:1–11CrossRefGoogle Scholar
  57. Ruiz MT, Voinnet O, Baulcombe DC (1998) Initiation and maintenance of virus-induced gene silencing. Plant Cell 10:937CrossRefGoogle Scholar
  58. Schmitz RJ, Schultz MD, Lewsey MG, O’Malley RC, Urich MA, Libiger O, Schork NJ, Ecker JR (2011) Transgenerational epigenetic instability is a source of novel methylation variants. Science 334:369–373CrossRefGoogle Scholar
  59. Sijen T, Vijn I, Rebocho A, van Blokland R, Roelofs D, Mol JNM, Kooter JM (2001) Transcriptional and posttranscriptional gene silencing are mechanistically related. Curr Biol 11:436–440CrossRefGoogle Scholar
  60. Song XW, Li PC, Zhai JX, Zhou M, Ma LJ, Liu B, Jeong DH, Nakano M, Cao SY, Liu CY, Chu CC, Wang XJ, Green PJ, Meyers BC, Cao XF (2012) Roles of DCL4 and DCL3b in rice phased small RNA biogenesis. Plant J 69:462–474CrossRefGoogle Scholar
  61. Stroud H, Ding B, Simon SA, Feng S, Bellizzi M, Pellegrini M, Wang G-L, Meyers BC, Jacobsen SE (2013) Plants regenerated from tissue culture contain stable epigenome changes in rice. eLife 2Google Scholar
  62. Thieme CJ, Rojas-Triana M, Stecyk E, Schudoma C, Zhang WN, Yang L, Minambres M, Walther D, Schulze WX, Paz-Ares J, Scheible WR, Kragler F (2016) Endogenous Arabidopsis messenger RNAs transported to distant tissues. Nat Plants 2:15025Google Scholar
  63. Tournier B, Tabler M, Kalantidis K (2006) Phloem flow strongly influences the systemic spread of silencing in GFP Nicotiana benthamiana plants. Plant J 47:383–394CrossRefGoogle Scholar
  64. Vaistij FE, Jones L (2009) Compromised virus-induced gene silencing in RDR6-deficient plants. Plant Physiol 149:1399–1407CrossRefGoogle Scholar
  65. Van der krol AR, Mur LA, Beld M, Mol JNM, Stuitje AR (1990) Flavonoid genes in petunia – addition of a limited number of gene copies may lead to a suppression of gene-expression. Plant Cell 2:291–299CrossRefGoogle Scholar
  66. Vazquez F, Vaucheret H, Rajagopalan R, Lepers C, Gasciolli V, Mallory A, Hilbert J, Bartel D, Crete P (2004) Endogenous trans-acting siRNAs regulate the accumulation of Arabidopsis mRNAs. Mol Cell 16:69–79CrossRefGoogle Scholar
  67. Vining K, Pomraning KR, Wilhelm LJ, Ma C, Pellegrini M, Di YM, Mockler TC, Freitag M, Strauss SH (2013) Methylome reorganization during in vitro dedifferentiation and regeneration of Populus trichocarpa. BMC Plant Biol 13:92CrossRefGoogle Scholar
  68. Wu L, Zhou H, Zhang Q, Zhang J, Ni F, Liu C, Qi Y (2010) DNA methylation mediated by a microRNA pathway. Mol Cell 38:465–475CrossRefGoogle Scholar
  69. Wu L, Mao L, Qi Y (2012) Roles of DICER-LIKE and ARGONAUTE proteins in TAS-derived small interfering RNA-triggered DNA methylation. Plant Physiol 160:990–999CrossRefGoogle Scholar
  70. Wu R, Wang X, Lin Y, Ma Y, Liu G, Yu X, Zhong S, Liu B (2013) Inter-species grafting caused extensive and heritable alterations of DNA methylation in Solanaceae plants. PLoS One 8:e61995CrossRefGoogle Scholar
  71. Zhang BH, Pan XP, Cannon CH, Cobb GP, Anderson TA (2006) Conservation and divergence of plant microRNA genes. Plant J 46:243–259CrossRefGoogle Scholar
  72. Zhang W, Kollwig G, Stecyk E, Apelt F, Dirks R, Kragler F (2014) Graft-transmissible movement of inverted-repeat-induced siRNA signals into flowers. Plant J 80:106–121CrossRefGoogle Scholar
  73. Zhong XH, Du JM, Hale CJ, Gallego-Bartolome J, Feng SH, Vashisht AA, Chory J, Wohlschlegel JA, Patel DJ, Jacobsen SE (2014) Molecular mechanism of action of plant DRM de novo DNA methyltransferases. Cell 157:1050–1060CrossRefGoogle Scholar

Copyright information

© Springer Nature Singapore Pte Ltd. 2018

Authors and Affiliations

  1. 1.College of Agriculture & BiotechnologyZhejiang UniversityZhejiangChina
  2. 2.Department of Agriculture and Life ScienceHirosaki UniversityHirosakiJapan

Personalised recommendations