Epigenetic maps of pearl millet reveal a prominent role for CHH methylation in regulating tissue-specific gene expression

Pearl millet (Pennisetum glaucum) is a major staple food in arid and semi-arid regions of sub-Saharan Africa, India, and South Asia. However, how epigenetic mechanisms regulate tissue-specific gene expression in this crop remains poorly understood. In this study, we profiled multiple epigenetic features in the young panicles and roots of pearl millet using RNA-seq, ATAC-seq, whole-genome bisulfite sequencing, and ChIP-seq (H3K4me3 and H3K36me3). We identified thousands of genes that were differentially expressed between these two tissues. Root-specific genes were enriched for plant hormone signaling, oxidative phosphorylation, and stress responses. Analysis of chromatin accessibility revealed that root-specific accessible chromatin regions (ACRs) were enriched in binding motifs for stress-responsive transcription factors (e.g., NAC, WRKY), whereas ACRs in young panicles were enriched in motifs for developmental regulators (e.g., AP2/ERF). DNA methylation profiling revealed 25,141 tissue-specific differentially methylated regions, with CHH methylation—rather than CG or CHG methylation—showing the strongest tissue specificity. Promoters of root-specific genes had higher levels of CHH methylation compared to those of young panicle–specific genes, suggesting that the roles of CHH methylation in regulating transcription might be tissue dependent. Notably, promoter-associated H3K4me3 marked panicle-specific genes, whereas root-specific expression was primarily linked to chromatin accessibility, suggesting a transcription factor–mediated regulatory mechanism. Together, our findings highlight the distinct epigenetic frameworks governing tissue-specific gene expression in pearl millet and provide valuable insights for advancing the genetic improvement of this crop.