Stable unmethylated DNA demarcates expressed genes and their cis-regulatory space in plant genomes

Abstract

The genomic sequences of crops continue to be produced at a frenetic pace. However, it remains challenging to develop complete annotations of functional genes and regulatory elements in these genomes. Here, we explore the potential to use DNA methylation profiles to develop more complete annotations. Using leaf tissue in maize, we define ∼100,000 unmethylated regions (UMRs) that account for 5.8% of the genome; 33,375 UMRs are found greater than 2 kilobase pairs from genes. UMRs are highly stable in multiple vegetative tissues and they capture the vast majority of accessible chromatin regions from leaf tissue. However, many UMRs are not accessible in leaf (leaf-iUMRs) and these represent a set of genomic regions with potential to become accessible in specific cell types or developmental stages. Leaf-iUMRs often occur near genes that are expressed in other tissues and are enriched for transcription factor (TF) binding sites of TFs that are also not expressed in leaf tissue. The leaf-iUMRs exhibit unique chromatin modification patterns and are enriched for chromatin interactions with nearby genes. The total UMRs space in four additional monocots ranges from 80-120 megabases, which is remarkably similar considering the range in genome size of 271 megabases to 4.8 gigabases. In summary, based on the profile from a single tissue, DNA methylation signatures pinpoint both accessible regions and regions poised to become accessible or expressed in other tissues. UMRs provide powerful filters to distill large genomes down to the small fraction of putative functional genes and regulatory elements.Significance StatementCrop genomes can be very large with many repetitive elements and pseudogenes. Distilling a genome down to the relatively small fraction of regions that are functionally valuable for trait variation can be like looking for needles in a haystack. The unmethylated regions in a genome are highly stable during vegetative development and can reveal the locations of potentially expressed genes or cis-regulatory elements. This approach provides a framework towards complete annotation of genes and discovery of cis-regulatory elements using methylation profiles from only a single tissue.

The maize gene maternal derepression of r1 (mdr1) encodes a DNA glycosylase that demethylates DNA and reduces siRNA expression in endosperm

“…Since imprinted gene expression in maize is also clearly linked to demethylation of maternal alleles (Zhang et al 2014), we expect that gametophyte defects are a consequence of mis-regulated gene expression rather than genome instability due to transposon activation. Such developmentally dynamic mCG and mCHG in and near genes would be exceptions to the general rule of such methylation being stable (Oka et al 2017;Crisp et al 2020). In multiple species, demethylation of transposons near MEGs is clearly connected to their maternal expression (Gehring et al 2009;Hatorangan et al 2016;Pignatta et al 2018;Rodrigues et al 2021).…”

Intergenic spaces: a new frontier to improving plant health

“…Given that gene expression depends on access to DNA, methods such as chromatin accessibility assays, nucleosome positioning, and surveillance of DNA epigenetic status enhance our understanding of genome dynamics and structural variations that influence gene regulation (Lu et al ., 2017, 2019; Gabur et al ., 2019; Crisp et al ., 2020). For example, transposase‐accessible chromatin using ‘ATAC‐seq’ enabled the discovery of thousands of previously unidentified CREs in diverse plant species, showing that, although sometimes found at distal locations, CREs are more typically enriched near genes (Lu et al ., 2019).…”

“…Other methods, such as DNA‐affinity purification sequencing (DAP‐seq), provide valuable information regarding transcription factor binding affinity and methylation sensitivity, and clarify the relevant transcriptional regulatory landscape of plants (O'Malley et al ., 2016). Combining chromatin accessibility with plant DNA methylation information is a powerful means to characterize complex genomes, and to identify new genes and their regulatory regions (Crisp et al ., 2020). Knowledge of relevant regulatory elements can be used to generate prediction models for gene expression variation due to stress responses, which can improve selection and breeding processes in any crop species (Meng et al ., 2021; Zhou et al ., 2021).…”

Genetic and epigenetic contributions to variation in transposable element expression responses to abiotic stress in maize

“…Unmethylated regions were determined following methods described in Crisp et al (2020). In brief, each 100bp tile was classified as missing data, RdDM, Heterochromatin, CG only, Unmethylated, or Intermediate according to their methylation state in each context.…”

“…We assessed whether there were factors that associated with the expression of portions of specific TEs. While TEs are generally highly methylated there are some unmethylated regions (UMRs) that occur within TEs (Crisp et al;Noshay et al, 2020). WGBS data was available for B73 control, cold and heat stress and was used to call UMRs ( Figure S6).…”