Genomas de plantas enxertadas "conversam" entre si

quarta-feira, janeiro 20, 2016

Mobile small RNAs regulate genome-wide DNA methylation

Mathew G. Lewsey a,b,1,2, Thomas J. Hardcastle c,1, Charles W. Melnyk c,3, Attila Molnar c,4, Adrián Valli c, Mark A. Urich a, Joseph R. Nery a, David C. Baulcombe c,5, and Joseph R. Ecker a,b,d,5

aGenomic Analysis Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037;

bPlant Biology Laboratory, The Salk Institute for Biological Studies, La Jolla, CA 92037;

cDepartment of Plant Sciences, University of Cambridge, Cambridge CB2 3EA, United Kingdom;

dHoward Hughes Medical Institute, The Salk Institute for Biological Studies, La Jolla, CA 92037

Edited by Steven E. Jacobsen, University of California, Los Angeles, CA, and approved December 15, 2015 (received for review July 29, 2015)

A graft between two genotypes of Arabidopsis thaliana, shown in a confocal microscopy image. One genotype has plasma membranes marked in yellow, and the other marked in red. Researchers studied the movement of sRNAs across the graft junction, and the resulting epigenetic changes in the plants’ genome.

Credit: Charles Melnyk at The Sainsbury Laboratory, Cambridge University


Small RNAs (sRNAs) of 24 nt are associated with transcriptional gene silencing by targeting DNA methylation to complementary sequences. We demonstrated previously that sRNAs move from shoot to root, where they regulate DNA methylation of three endogenous transposable elements (TEs). However, the full extent of root DNA methylation dependent on mobile sRNAs was unknown. We demonstrate that DNA methylation at thousands of sites depends upon mobile sRNAs. These sites are associated with TE superfamilies found in gene-rich regions of the genome, which lose methylation selectively in an sRNA-deficient mutant. If the TEs were able to reactivate, they could cause genome instability and altered gene expression patterns, with negative effects on the plant. Consequently, mobile sRNAs may defend against these TEs.

RNA silencing at the transcriptional and posttranscriptional levels regulates endogenous gene expression, controls invading transposable elements (TEs), and protects the cell against viruses. Key components of the mechanism are small RNAs (sRNAs) of 21–24 nt that guide the silencing machinery to their nucleic acid targets in a nucleotide sequence-specific manner. Transcriptional gene silencing is associated with 24-nt sRNAs and RNA-directed DNA methylation (RdDM) at cytosine residues in three DNA sequence contexts (CG, CHG, and CHH). We previously demonstrated that 24-nt sRNAs are mobile from shoot to root in Arabidopsis thaliana and confirmed that they mediate DNA methylation at three sites in recipient cells. In this study, we extend this finding by demonstrating that RdDM of thousands of loci in root tissues is dependent upon mobile sRNAs from the shoot and that mobile sRNA-dependent DNA methylation occurs predominantly in non-CG contexts. Mobile sRNA-dependent non-CG methylation is largely dependent on the DOMAINS REARRANGED METHYLTRANSFERASES 1/2 (DRM1/DRM2) RdDM pathway but is independent of the CHROMOMETHYLASE (CMT)2/3 DNA methyltransferases. Specific superfamilies of TEs, including those typically found in gene-rich euchromatic regions, lose DNA methylation in a mutant lacking 22- to 24-nt sRNAs (dicer-like 2, 3, 4 triple mutant). Transcriptome analyses identified a small number of genes whose expression in roots is associated with mobile sRNAs and connected to DNA methylation directly or indirectly. Finally, we demonstrate that sRNAs from shoots of one accession move across a graft union and target DNA methylation de novo at normally unmethylated sites in the genomes of root cells from a different accession.

RNA-directed DNA methylation plant grafting transposable element small RNA transcriptional gene silencing


1M.G.L. and T.J.H. contributed equally to this work.

2Present address: Centre for AgriBioscience, Department of Animal, Plant and Soil Science, School of Life Science, La Trobe University, Bundoora, VIC 3086, Australia.

3Present address: Sainsbury Laboratory, University of Cambridge, Cambridge CB2 1LR, United Kingdom.

4Present address: Institute of Molecular Plant Sciences, School of Biological Sciences, The University of Edinburgh, Edinburgh EH9 3JR, United Kingdom.

5To whom correspondence may be addressed. Email: dcb40{at} or ecker{at}

Author contributions: M.G.L., T.J.H., C.W.M., A.M., D.C.B., and J.R.E. designed research; M.G.L., T.J.H., C.W.M., A.M., A.V., M.A.U., and J.R.N. performed research; T.J.H. contributed new analytic tools; M.G.L., T.J.H., C.W.M., A.M., D.C.B., and J.R.E. analyzed data; and M.G.L., T.J.H., C.W.M., A.M., D.C.B., and J.R.E. wrote the paper.

The authors declare no conflict of interest.

This article is a PNAS Direct Submission.

Data deposition: The data reported in this paper have been deposited in the European Nucleotide Archive (accession no. E-MTAB-3473).

This article contains supporting information online at

Freely available online through the PNAS open access option.