DNA methylation around transcription start sites is not globally associated with transcription in the grain of natural and synthetic hexaploid wheat

Epigenetic mechanisms including DNA methylation are assumed to play crucial roles in the maintenance of genome integrity, regulation of gene expression and development, and their increasing exploitation in breeding applications is anticipated. However, the relationship between DNA methylation and gene expression remains ambiguous and difficult to generalize. Here we explored the hypothesized causality between the level of transcription and cytosine methylation at the 5’ end of genes (around transcription start sites and start codons) in relation to whole-genome duplication in natural and synthetic allohexaploid wheat (Triticum/Aegilops complex). Using transcriptomes and a sequence capture protocol coupled with bisulfite sequencing, we observed sometimes significant, but overall very weak associations between gene expression and 5’ end methylation on a genome-wide scale. In synthetic wheat allohexaploids, global methylation differences between subgenomes are not triggered by the polyploidization, as the subgenome patterns are rather faithfully inherited from parents. A small number of genes differentially methylated between the parents and synthetics was consistently recovered in reciprocal synthetics and subsequent generations. Differences in transcription between homeologs are not clearly associated with 5’ end methylation in either natural or synthetic wheat. Overall, allopolyploidization triggers only minor methylation changes around transcription start sites and start codons of nascent wheat allopolyploids, and these are not statistically associated with differential expression. Although there is a measurable methylation difference between silent and expressed genes in the developing grain, our results do not support the hypothesis that 5’ end DNA methylation is engaged in the regulation of gene expression in natural and synthetic wheat. While a ‘genome shock’ hypothesis predicts extensive transcriptomic and epigenetic reorganization after polyploidization, DNA methylation patterns around transcription start sites are generally undisturbed in nascent wheat allohexaploids. Although this stability might indicate importance for gene regulation, a clear relationship between DNA methylation and transcription was not observed either on a genome-wide scale, or among triads of homeologous genes.