Understanding epigenetic regulation in non-model plants: Transcriptomic responses to seed demethylation in leaves and roots of the annual herb Erodium cicutarium (Geraniaceae)

Abstract

Epigenetic regulation has emerged as a significant element in adaptation to heterogeneous and stressful environments, with modifications in DNA methylation being particularly relevant in plants. DNA methylation inhibitors have been used to investigate the relationship between DNA methylation and plastic plant phenotypes. However, their effect in gene expression regulation along lifetime remains understudied in non-model plants. Here, we analyze the effects of seed exposure to 5-azacytidine (5-azaC) in plant gene regulation. Scarified seeds from a single inbred line of Erodium cicutarium were soaked for 48 h in either water or a low concentration solution of 5-azaC before sowing. Subsequently, RNA was extracted from juvenile roots, juvenile leaves and adult leaves, and their transcriptomes were sequenced. Differential gene expression analysis was performed between treatments (control vs. treated) for all tissues together and separately. Beforehand, a draft genome of E. cicutarium was assembled to use it as reference for the transcriptome analysis, and its DNA methyltransferase genes were characterized. We found that 5-azaC up-regulated chromomethylase CMT1 across all treated samples, and the domain rearranged DNA methyltransferase DRM2 in juvenile roots. Furthermore, adult leaves showed more differentially expressed genes between control and 5-azaC treated samples compared to juvenile leaves, supporting long term transcriptomic effects of a short exposure to 5-azaC at seed germination. At adult stage, leaves of individuals treated with 5-azaC exhibited up-regulation of genes involved in seed dormancy release, photoinhibition, and osmotic stress. Finally, gene co-expression network analysis revealed a module of co-expressed genes with differential gene expression linked to 5-azaC treatment in juvenile roots, that was enriched with genes involved in retrotransposon activity and in anthocyanin metabolism. Altogether, this study illustrates how the experimental treatment with 5-azaC at seed stage generates tissue- and age-specific transcriptional shifts, directly affecting gene regulation and potentially broadening phenotype variation in this fast-growing annual plant.