Increase of DNA Methylation at the HvCKX2.1 Promoter by Terminal Drought Stress in Barley

Abstract

Terminal drought stress during grain filling is the major abiotic factor that limits crop yield in barley. The mother plant acclimates to the environment and perceives signals that result in a change of the physiological state within the grain and therefore affect the seed development and germination of the next generation. Small regulatory RNAs have been described to be involved in plant drought stress response by suppressing the respective target genes. Based on their origin and function, these small RNAs are classified as micro RNAs (miRNA), short interfering RNAs (siRNA) or heterochromatic small interfering RNA (hc-siRNA). In addition, 24mer sized hc-siRNAs are associated with RNA directed DNA methylation (RdDM) and transcriptional gene silencing (TGS). The analysis of hc-siRNA by small RNA sequencing in barley caryopses after imposition of terminal drought stress allowed the identification of stress specific 24mers. Based on the sequence homology of the siRNAs to the promoter region of CYTOKININ-OXIDASE 2.1 (HvCKX2.1), this putative target gene was selected for further investigation. Terminal drought stress leads to an increased level of DNA methylation at the HvCKX2.1 promoter and the seeds derived from drought stressed plants showed faster shoot emergence. Accumulation of cytokinin ribosides, which are the known substrates of cytokinin-oxidase, can explain the observed phenotype of faster shoot emergence from seeds of drought stressed mother plants. Analysis of transgenic plants with modulated levels of abscisic acid (ABA) in the grain confirmed the ABA/drought stress responsive ProHvCKX2.1 methylation and correlation with shoot emergence speed.

Epigenetic responses to abiotic stresses during reproductive development in cereals

“…A global DNA methylation study during seed development in rice showed that the expression of genes responding to the growth hormone auxin and the stress hormone ABA was controlled by DNA methylation (Xing et al 2015 ). Recently, this has also been described in barley (Surdonja et al 2017 ), indicating that the cellularization process of cereal endosperm could be especially stress-sensitive. Therefore, it was suggested that DNA methylation controls ABA levels and thus promotes endosperm cellularization in cereals.…”

“…In barley and wheat, AGO proteins, which participate in gene silencing through RdDM pathways and which are components of the RNA-induced silencing complex (RISC), were shown to regulate seed maturation and dormancy (Singh and Singh 2012 ; Singh et al 2014 ). When barley plants were submitted to terminal stress during grain filling, increased DNA methylation was observed, for example, in the promoter region of the Cytokinin - Oxidase 2.1 ( CKX2.1 ) gene, which is a target of the RdDM pathway (Surdonja et al 2017 ). Moreover, it was reported that dormancy was reduced at both increased temperatures and water deficit during seed maturation (Begcy and Walia 2015b ; Singh et al 2014 ).…”

Epigenetics: possible applications in climate-smart crop breeding

“…Terminal drought stress during grain filling is the major abiotic factor that limits crop yield in barley. Stress-specific 24mer size hc-siRNA was found in the promoter regions of the barley cytokinin-oxidase 2.1 gene ( HvCKX2.1 ) in the caryopsis exposed to terminal drought ( Surdonja et al , 2017 ). The authors found that under terminal drought the level of DNA methylation of this gene was increased.…”

“…This mechanism was found to be involved in the drought stress response in barley in which in drought-stressed plants the promoter region of cytokinin-oxidase 2.1 ( HvCKX 2.1 ) had an increased level of DNA methylation. Furthermore, the seeds from the drought-stressed plants showed faster shoot emergence due to an abundance of cytokinin ribosides ( Surdonja et al , 2017 ). In rice, it was found that a high proportion of the drought-induced epimutations (DNA methylation changes) maintained their altered methylation pattern in successive generations exposed to drought from the tillering to the grain filling stage, suggesting the presence of possible epi-marks that are drought inducible and heritable across generations ( Zheng et al , 2017 ).…”

Induced Methylation in Plants as a Crop Improvement Tool: Progress and Perspectives

“…Accurate DNA methylation mark induction and removal have complementary functions and could both play a role in crop improvement. DNA hypermethylation is, for example, involved in crop stress resistance mechanisms, such as wheat salt tolerance, pea and barley response to drought stress, and tomato response to cold [58][59][60][61]. Meanwhile, DNA hypomethylation is implicated in drought stress response in rice and faba beans, and heat response in soybean and rapeseed [62][63][64][65].…”