Genomic regions free of nucleosomes, which are hypersensitive to DNase I digestion, are known as DNase I hypersensitive sites (DHSs) and frequently contain cis-regulatory DNA elements. To investigate their prevalence and characteristics in maize (), we developed high-resolution genome-wide DHS maps using a modified DNase-seq technique. Maize DHSs exhibit depletion of nucleosomes and low levels of DNA methylation and are enriched with conserved noncoding sequences (CNSs). We developed a protoplast-based transient transformation assay to assess the potential gene expression enhancer and/or promoter functions associated with DHSs, which showed that more than 80% of DHSs overlapping with CNSs showed an enhancer function. Strikingly, nearly 25% of maize DHSs were derived from transposable elements (TEs), including both class I and class II transposons. Interestingly, TE-derived DHSs (teDHSs) homologous to retrotransposons were enriched with sequences related to the intrinsic cis-regulatory elements within the long terminal repeats of retrotransposons. We demonstrate that more than 80% of teDHSs can drive transcription of a reporter gene in protoplast assays. These results reveal the widespread occurrence of TE-derived cis-regulatory sequences and suggest that teDHSs play a major role in transcriptional regulation in maize.

Recombination plays an integral role in the creation of novel genetic variation in sexually reproducing species. Despite this important role, the determinants and evolution of crossover hotspots have remained poorly understood in plants. Here, we present a comparative analysis of two rice (Oryza sativa) historical recombination maps from two subspecies (indica and japonica) using 150 resequenced genomes. Fine-scale recombination rates and crossover hotspots were validated by comparison with a consensus genetic map and empirically derived crossovers, respectively. Strikingly, nearly 80% of crossover hotspots were unique to each subspecies, despite their relatively recent divergence and broad-scale correlated recombination rates. Crossover hotspots were enriched with Stowaway and P instability factor (PIF)/Harbinger transposons and overlapped accessible chromatin regions. Increased nucleotide diversity and signatures of population differentiation augmented by Stowaway and PIF/Harbinger transposons were prevalent at subspecies-specific crossover hotspots. Motifs derived from lineage-specific indica and japonica crossover hotspots were nearly identical in the two subspecies, implicating a core set of crossover motifs in rice. Finally, Stowaway and PIF/Harbinger transposons were associated with stabilized G/C bias within highly active hotspots, suggesting that hotspot activity can be fueled by de novo variation. These results provide evolutionary insight into historical crossover hotspots as potentially powerful drivers of sequence and subspecies evolution in plants.

Summary Although polyploid plants have larger leaves than their diploid counterparts, the molecular mechanisms underlying this difference (or trait) remain elusive. Differentially expressed genes (DEGs) between triploid and full‐sib diploid poplar trees were identified from two transcriptomic data sets followed by a gene association study among DEGs to identify key leaf growth regulators. Yeast one‐hybrid system, electrophoretic mobility shift assay, and dual‐luciferase assay were employed to substantiate that PpnGRF5‐1 directly regulated PpnCKX1. The interactions between PpnGRF5‐1 and growth‐regulating factor (GRF)‐interacting factors (GIFs) were experimentally validated and a multilayered hierarchical regulatory network (ML‐hGRN)‐mediated by PpnGRF5‐1 was constructed with top‐down graphic Gaussian model (GGM) algorithm by combining RNA‐sequencing data from its overexpression lines and DAP‐sequencing data. PpnGRF5‐1 is a negative regulator of PpnCKX1. Overexpression of PpnGRF5‐1 in diploid transgenic lines resulted in larger leaves resembling those of triploids, and significantly increased zeatin and isopentenyladenine in the apical buds and third leaves. PpnGRF5‐1 also interacted with GIFs to increase its regulatory diversity and capacity. An ML‐hGRN‐mediated by PpnGRF5‐1 was obtained and could largely elucidate larger leaves. PpnGRF5‐1 and the ML‐hGRN‐mediated by PpnGRF5‐1 were underlying the leaf growth and development.

The elucidation of epigenetic responses of salt-responsive genes facilitates understanding of the underlying mechanisms that confer salt tolerance in rice. However, it is still largely unknown how epigenetic mechanisms are associated with the expression of salt-responsive genes in rice and other crops. In this study, we reported tissue-specific gene expression and tissue-specific changes in chromatin modifications or signatures between seedlings and roots in response to salt treatment. Our study indicated that among six of individual mark examined (H3K4me3, H3K27me3, H4K12ac, H3K9ac, H3K27ac and H3K36me3), a positive association between salt-related changes in histone marks and the expression of differentially expressed genes (DEGs) was observed only for H3K9ac and H4K12ac in seedlings and H3K36me3 in roots. In contrast, chromatin states (CSs) with combinations of six histone modification marks played crucial roles in the differential expression of salt-responsive genes between seedlings and roots. Most importantly, CS7 containing the bivalent marks H3K4me3 and H3K27me3, with a mutual exclusion of functions with each other, displayed distinct functions in the expression of DEGs in both tissues. Specifically, H3K27me3 in CS7 mainly suppressed the expression of DEGs in roots, while H3K4me3 affected the expression of down- and up-regulated genes, possibly by antagonizing the repressive role of H3K27me3 in seedlings. Our findings indicate distinct impacts of the CSs on the differential expression of salt-responsive genes between seedlings and roots in rice, which provides an important background for understanding chromatin-based epigenetic mechanisms that might confer salt tolerance in plants.

Background: Maize mesophyll (M) cells play important roles in various biological processes such as photosynthesis II and secondary metabolism. Functional differentiation occurs during M-cell development, but the underlying mechanisms for regulating M-cell development are largely unknown. Results: We conducted single-cell RNA sequencing (scRNA-seq) to profile transcripts in maize leaves. We then identified coregulated modules by analyzing the resulting pseudo-time-series data through gene regulatory network analyses. WRKY, ERF, NAC, MYB and Heat stress transcription factor (HSF) families were highly expressed in the early stage, whereas CONSTANS (CO)-like (COL) and ERF families were highly expressed in the late stage of M-cell development. Construction of regulatory networks revealed that these transcript factor (TF) families, especially HSF and COL, were the major players in the early and later stages of M-cell development, respectively. Integration of scRNA expression matrix with TF ChIP-seq and Hi-C further revealed regulatory interactions between these TFs and their targets. HSF1 and COL8 were primarily expressed in the leaf bases and tips, respectively, and their targets were validated with protoplast-based ChIP-qPCR, with the binding sites of HSF1 being experimentally confirmed. Conclusions: Our study provides evidence that several TF families, with the involvement of epigenetic regulation, play vital roles in the regulation of M-cell development in maize.

Summary Transposons significantly contribute to genome fractions in many plants. Although numerous transposon‐related mutations have been identified, the evidence regarding transposon‐derived genes regulating crop yield and other agronomic traits is very limited. In this study, we characterized a rice Harbinger transposon‐derived gene called PANICLE NUMBER AND GRAIN SIZE (PANDA), which epigenetically coordinates panicle number and grain size. Mutation of PANDA caused reduced panicle number but increased grain size in rice, while transgenic plants overexpressing this gene showed the opposite phenotypic change. The PANDA‐encoding protein can bind to the core polycomb repressive complex 2 (PRC2) components OsMSI1 and OsFIE2, and regulates the deposition of H3K27me3 in the target genes, thereby epigenetically repressing their expression. Among the target genes, both OsMADS55 and OsEMF1 were negative regulators of panicle number but positive regulators of grain size, partly explaining the involvement of PANDA in balancing panicle number and grain size. Moreover, moderate overexpression of PANDA driven by its own promoter in the indica rice cultivar can increase grain yield. Thus, our findings present a novel insight into the epigenetic control of rice yield traits by a Harbinger transposon‐derived gene and provide its potential application for rice yield improvement.

SERRATE (SE) is a core protein for microRNA (miRNA) biogenesis as well as for mRNA alternative splicing. Investigating the regulatory mechanism of SE expression is hence critical to understanding its detailed function in diverse biological processes. However, little about the control of SE expression has been clarified, especially through long noncoding RNA (lncRNA). Here, we identified an antisense intragenic lncRNA transcribed from the 3′ end of SE , named SEAIRa. SEAIRa repressed SE expression, which in turn led to serrated leaves. SEAIRa recruited plant U-box proteins PUB25/26 with unreported RNA binding ability and a ubiquitin-like protein related to ubiquitin 1 (RUB1) for H2A monoubiquitination (H2Aub) at exon 11 of SE . In addition, PUB25/26 helped cleave SEAIRa and release the 5′ domain fragment, which recruited the PRC2 complex for H3 lysine 27 trimethylation (H3K27me3) deposition at the first exon of SE . The distinct modifications of H2Aub and H3K27me3 at different sites of the SE locus cooperatively suppressed SE expression. Collectively, our results uncover an epigenetic mechanism mediated by the lncRNA SEAIRa that modulates SE expression, which is indispensable for plant growth and development.

The success of common wheat as a global staple crop was largely attributed to its genomic diversity and redundancy due to the merge of different genomes, giving rise to the major question how subgenome-divergent and -convergent transcription is mediated and harmonized in a single cell. Here, we create a catalog of genome-wide transcription factor-binding sites (TFBSs) to assemble a common wheat regulatory network on an unprecedented scale. A significant proportion of subgenome-divergent TFBSs are derived from differential expansions of particular transposable elements (TEs) in diploid progenitors, which contribute to subgenome-divergent transcription. Whereas subgenome-convergent transcription is associated with balanced TF binding at loci derived from TE expansions before diploid divergence. These TFBSs have retained in parallel during evolution of each diploid, despite extensive unbalanced turnover of the flanking TEs. Thus, the differential evolutionary selection of paleo- and neo-TEs contribute to subgenome-convergent and -divergent regulation in common wheat, highlighting the influence of TE repertory plasticity on transcriptional plasticity in polyploid.