In total, approximately 10,000 regions of the genome were determined as differentially accessible regions (DARs) upon induction of bZIP11 translocation to the nucleus, depending on which control was used (Table 1). Approximately 50,000 cells from each replicate of each treatment were used for ATAC-seq. In parallel, the samples were treated, prior to DEX or acetone treatment, with cycloheximide (CHX), a protein biosynthesis inhibitor 21 that enables to discriminate the direct targets of bZIP11 from its indirect targets. Using protoplasts allows for a rapid and homogeneous chemical uptake which reduces variability between samples 20. Four biological replicates were treated with either DEX, which induces the translocation of bZIP11 protein into the nucleus, or acetone, the solvent for DEX (Mock control). Protoplasts were treated for 45 minutes after a one hr recovery period following protoplast extraction. Four-week-old Arabidopsis plants were used to extract leaf protoplasts from approximately six leaves per plant (experiment summarised in Fig. Upon treatment with dexamethasone (DEX), which binds to the HBD, the translocation of bZIP11 to the nucleus is enabled, allowing its TF activity to occur 19. A rat glucocorticoid receptor, or hormone binding domain (HBD), is fused to bZIP11, and retains this TF in the cytoplasm. In these p35S:bZIP11-HBD plants, the expression of bZIP11 coding sequence is driven by the cauliflower mosaic virus, CaMV35S, promoter ( p35S) that leads to ectopic gene expression in the whole plant. To achieve a comprehensive profile of accessible chromatin regions and discover new downstream targets of bZIP11, we generated 24 chromatin accessibility data sets from wild-type (WT) Arabidopsis protoplast samples, as well as from transgenic p35S:bZIP11-HBD Arabidopsis plants 18. Here we report ATAC-seq results in response to induction of a single plant TF, with known regulatory sequences, to determine the genome-wide effect of this TF on chromatin accessibility in Arabidopsis. Previous studies have performed ATAC-seq on different plant organs, conditions, or in response to different treatments to determine changes in accessibility of those DNA regions that are likely to contain regulatory sequences 14, 15, 16, 17. This technique is rapid and can be performed on small quantities of starting material. However, the extent to which bZIP11 impacts chromatin accessibility at the whole-genome scale remains unknown.Īssay for Transposase-Accessible Chromatin with high throughput sequencing (ATAC-seq) is a technique which can determine genome-wide locations of chromatin accessible to transposase insertion and by extension, other proteins, for example TFs 12, 13. bZIP11 has been demonstrated to promote the expression of specific auxin-related genes by recruiting histone acetylation machinery to enhance chromatin accessibility 3, 11. The bZIP TFs bind DNA at cis-regulatory elements (CREs) known as G-boxes, which all have a conserved ACGT core flanked by cytosines or guanines (C/GACGTG/C) 10. It is estimated by DNA Affinity Purification with high throughput sequencing (DAP-seq) that bZIP11 contains DNA-binding sites in over 7,000 genes in Arabidopsis, which is nearly one third of the entire genes in the genome 9. In Arabidopsis thaliana (Arabidopsis), bZIP11 belongs to a group of five proteins involved in sugar responses, named the S 1 bZIP group 2, 4, 5, 6, 7, 8. Our study provides a resource for understanding how bZIP11 regulates the genome at the chromatin level and provides an example of the impact of a single transcription factor on a whole plant genome.īasic leucine zipper11 (bZIP11) is a transcription factor (TF) which regulates gene expression during low-energy conditions in plants and enables plants to adjust their metabolism, growth, and development to such unfavourable conditions 1, 2, 3. We identified, on average, 10,000 differentially accessible regions upon bZIP11 induction, corresponding to over 8,420 different genes out of the 25,000 genes present in the Arabidopsis genome. Here we addressed this by performing an ATAC-seq (Assay for Transposase-Accessible Chromatin with high-throughput sequencing) on Arabidopsis thaliana (Arabidopsis) leaf protoplasts to obtain a profile of chromatin patterning in response upon bZIP11 induction. Although previous results indicate that bZIP11 regulates chromatin accessibility based on evidence obtained from single genomic loci, to what extent this transcription factor regulates the chromatin landscape at the whole genome level remains unknown. Basic leucine zipper 11 (bZIP11) is a transcription factor that is activated under low energy conditions in plants and plays a crucial role in enabling plants to adapt to starvation situations.
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