Alterations of lysine modifications on the histone H3 N-tail under drought stress conditions in Arabidopsis thaliana

Post-translational modification of histone N-tails affects eukaryotic gene activity. In Arabidopsis, the histone modification level correlates with gene activation and repression in vernalization and flowering processes, but there is little information on changes in histone modification status and nucleosome structure under abiotic stresses. We determined the temporal and spatial changes in nucleosome occupancy and levels of H3K4me3, H3K9ac, H3K14ac, H3K23ac and H3K27ac in the histone H3 N-tail on the regions of four Arabidopsis drought stress-inducible genes, RD29A, RD29B, RD20 and At2g20880 [corrected], under drought stress conditions by chromatin immunoprecipitation analysis. We found two types of regulatory mechanisms of nucleosome occupancy function in the drought stress response. For RD29A and RD29B genes, nucleosome occupancy of promoter regions is low compared with that of coding regions, and no notable nucleosome loss occurs under drought stress. In contrast, nucleosome density is gradually decreased in response to drought stress on RD20 and At2g20880 [corrected] genes. Enrichments of H3K4me3 and H3K9ac correlate with gene activation in response to drought stress in all four genes. Interestingly, establishment of H3K4me3 occurs after accumulation of RNAPII on the coding regions of RD29A and At2g20880 [corrected]. Enrichment of H3K23ac and H3K27ac occurs in response to drought stress on the coding regions of RD29B, RD20 and At2g20880 [corrected], but not on the coding region of At2g20880 [corrected]. Our results indicate that histone modifications on the H3 N-tail are altered with gene activation on the coding regions of drought stress-responsive genes under drought stress conditions and that several patterns of nucleosome changes function in the drought stress response.     

Natural variation of H3K27me3 distribution between two Arabidopsis accessions and its association with flanking transposable elements

Background

Histone H3 lysine 27 tri-methylation and lysine 9 di-methylation are independent repressive chromatin modifications in Arabidopsis thaliana. H3K27me3 is established and maintained by Polycomb repressive complexes whereas H3K9me2 is catalyzed by SUVH histone methyltransferases. Both modifications can spread to flanking regions after initialization and were shown to be mutually exclusive in Arabidopsis.

Results

We analyzed the extent of natural variation of H3K27me3 in the two accessions Landsberg erecta (Ler) and Columbia (Col) and their F1 hybrids. The majority of H3K27me3 target genes in Col were unchanged in Ler and F1 hybrids. A small number of Ler-specific targets were detected and confirmed. Consistent with a cis-regulatory mechanism for establishing H3K27me3, differential targets showed allele-specific H3K27me3 in hybrids. Five Ler-specific targets showed the active mark H3K4me3 in Col and for this group, differential H3K27me3 enrichment accorded to expression variation. On the other hand, the majority of Ler-specific targets were not expressed in Col, Ler or 17 other accessions. Instead of H3K27me3, the antagonistic mark H3K9me2 and other heterochromatic features were observed at these loci in Col. These loci were frequently flanked by transposable elements, which were often missing in the Ler genome assembly.

Conclusion

There is little variation in H3K27me3 occupancy within the species, although H3K27me3 targets were previously shown as overrepresented among differentially expressed genes. The existing variation in H3K27me3 seems mostly explained by flanking polymorphic transposable elements. These could nucleate heterochromatin, which then spreads into neighboring H3K27me3 genes, thus converting them to H3K9me2 targets.     

Immune Mediators Regulate CFTR Expression through a Bifunctional Airway-Selective Enhancer

An airway-selective DNase-hypersensitive site (DHS) at kb −35 (DHS-35kb) 5′ to the cystic fibrosis transmembrane conductance regulator (CFTR) gene is evident in many lung cell lines and primary human tracheal epithelial cells but is absent from intestinal epithelia. The DHS-35kb contains an element with enhancer activity in 16HBE14o- airway epithelial cells and is enriched for monomethylated H3K4 histones (H3K4me1). We now define a 350-bp region within DHS-35kb which has full enhancer activity and binds interferon regulatory factor 1 (IRF1) and nuclear factor Y (NF-Y) in vitro and in vivo. Small interfering RNA (siRNA)-mediated depletion of IRF1 or overexpression of IRF2, an antagonist of IRF1, reduces CFTR expression in 16HBE14o- cells. NF-Y is critical for maintenance of H3K4me1 enrichment at DHS-35kb since depletion of NF-YA, a subunit of NF-Y, reduces H3K4me1 enrichment at this site. Moreover, depletion of SETD7, an H3K4 monomethyltransferase, reduces both H3K4me1 and NF-Y occupancy, suggesting a requirement of H3K4me1 for NF-Y binding. NF-Y depletion also represses Sin3A and reduces its occupancy across the CFTR locus, which is accompanied by an increase in p300 enrichment at multiple sites. Our results reveal that the DHS-35kb airway-selective enhancer element plays a pivotal role in regulation of CFTR expression by two independent regulatory mechanisms.