Small RNA and epigenetic silencing of transposable elements in grapevine embryogenic cell cultures

Abstract

Transposable elements (TEs) are a ubiquitous mutagenic force within the genomes of eukaryotes. Their mobilisation can rearrange gene exon sequences and regulatory motifs, leading to altered phenotypes. To prevent this potentially deleterious activity, transposon activity is usually highly repressed, particularly in the gametes, by localised epigenetic modification of the DNA. Grapevine is a unique and interesting model in which to study the activity of endogenous TEs since it has been vegetatively propagated for thousands of years. As a result, an accumulation of somatic mutations defines the genotypes of elite clones currently farmed. In order to study the transposition rates, insertion bias an impacts of endogenous TEs in grapevine, we have been regenerating vines from embryogenic callus cells. We have found that through this process, transposon activity is increased, allowing the recovery of a population of new clonal diversity. To characterise the biology of this process, we have completed the first whole-methylome analysis of grapevine tissue. Our results show that much of the conserved epigenetic silencing of TEs seen in vegetative cells is lost in totipotent embryogenic callus cultures, particularly for those TEs that lie within genes. Using massively parallel sequencing of small RNAs, which guide the enzyme complexes that methylate DNA, we observed an increased targeting of TEs for repression in embryogenic callus. This was accompanied by an abundance of new methylation around transposable elements independent of their context, indicative of trans-silencing. Although the consequences of historical TE activity is well-studied, the data presented here reveal the real-time process by which dormant transposons become active, create new mutations, and are once again silenced by their host cells in a wild-type genome. This provides insight into the process of somatic mutation and consequent clonal diversification.