Epigenetic differences between transgenes and endogenous genes promote transgene silencing in plants
Transgene loci that spontaneously undergo RNA silencing have been instrumental to decipher RNA silencing pathways in plants. In contrast, transgene loci that are stably expressed have been poorly characterized. Here we show that stably expressed transgene loci epigenetically differ from endogenous genes. Impairing the histone H3K4me3 demethylase JMJ14 increases DNA methylation at stably expressed transgene loci but not at endogenous genes, whereas impairing the histone H3K9me2 demethylase IBM1 increases DNA methylation at endogenous genes, but decreases DNA methylation at stably expressed transgene loci. Moreover, impairing IBM1 promotes post-transcriptional gene silencing (PTGS) at stably expressed transgene loci, whereas impairing JMJ14 prevents PTGS induced systemically by grafting onto silenced rootstocks. Given that impairing DNA methylation suppresses the effect of impairing JMJ14 and restores transgene capacity to undergo graft-induced PTGS, we propose that transgene DNA methylation prevents PTGS, likely by limiting the transcription of aberrant RNA by-products that are transformed into dsRNA by cellular RNA-dependent RNA polymerases to activate PTGS. Overall design: Genomic DNA from leaf samples analyzed by whole-genome bisulfite sequencing. Experiment 1: Leaf samples from 5-week-old plants carrying the 6b4 transgene in wild type Col-0 (6b4-1) and in mutants (6b4 jmj14, 6b4 jmj14 drm2 cmt3 and 6b4 drm2 cmt3). Experiment 2: Leaf samples of 8-week-old plants carrying the 6b4 transgene in wild type Col-0 (6b4-2) and mutants (6b4 nac50 nac52 and 6b4 ibm1).