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RNA-directed DNA methylation efficiency depends on trigger and target sequence identity.

Athanasios Dalakouras1, Elena Dadami1, Michèle Wassenegger1

  • 1RLP AgroScience GmbH, AlPlanta-Institute for Plant Research, Neustadt, 67435, Germany.

The Plant Journal : for Cell and Molecular Biology
|April 29, 2016
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Summary

RNA-directed DNA methylation (RdDM) in plants is guided by RNA molecules. This study shows RdDM efficiency depends on RNA sequence identity, not just small interfering RNA (siRNA) presence, suggesting longer RNAs may trigger RdDM.

Keywords:
DNA methylationRNA silencingbisulfite sequencingsmall RNAsviroids

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Area of Science:

  • Plant molecular biology
  • Epigenetics
  • RNA biology

Background:

  • RNA-directed DNA methylation (RdDM) is a key epigenetic mechanism in plants.
  • The precise nature of RNA molecules guiding RdDM and their target complementarity requirements remain incompletely understood.

Purpose of the Study:

  • To investigate the role of RNA sequence identity in guiding RNA-directed DNA methylation (RdDM).
  • To determine if small interfering RNAs (siRNAs) are the sole triggers for RdDM.
  • To explore the potential of longer RNAs in triggering RdDM.

Main Methods:

  • Generation of transgenic Nicotiana tabacum (Nt) plants expressing homologous and partially homologous viroid transgenes.
  • Genetic crossing of transgenic lines to create progeny with both transgenes.
  • Deep sequencing of small RNAs (sRNAs) and bisulfite sequencing to analyze methylation patterns and siRNA distribution.

Main Results:

  • TASVd-derived siRNAs were abundant and targeted both homologous TASVd and partially homologous SB2 transgenes.
  • Dense cis-RdDM occurred on the TASVd transgene, while weaker, heterogeneous trans-RdDM occurred on the SB2 transgene.
  • Trans-RdDM on SB2 was observed even without detectable homologous siRNAs, indicating RdDM is not solely dependent on siRNA occupancy.

Conclusions:

  • RdDM efficiency is positively correlated with trigger-target sequence identity.
  • RdDM can be triggered by factors beyond siRNA occupancy, potentially involving longer RNAs (>24 nt).
  • These findings advance our understanding of the specificity and mechanisms of RNA-directed DNA methylation in plants.