Distinct guard cell specific remodeling of chromatin accessibility during abscisic acid and CO2 dependent stomatal regulation (ATAC-Seq)

In plants, epidermal guard cells integrate and respond to numerous environmental signals to control stomatal pore apertures thereby regulating gas exchange. Chromatin structure controls transcription factor access to the genome, but whether large-scale chromatin remodeling occurs in guard cells during stomatal movements, and in response to the hormone abscisic acid (ABA) in general, remain unknown. Here we isolate guard cell nuclei from Arabidopsis thaliana plants to examine whether the physiological signals, ABA and CO2, regulate guard cell chromatin during stomatal movements. Our cell type specific analyses uncover patterns of chromatin accessibility specific to guard cells and define novel cis-regulatory sequences supporting guard cell specific gene expression. We find that ABA triggers extensive and dynamic chromatin remodeling in guard cells, roots, and mesophyll cells with clear patterns of cell-type specificity. DNA motif analyses uncover binding sites for distinct transcription factors enriched in ABA-induced and ABA-repressed chromatin. We identify the ABF/AREB bZIP-type transcription factors that are required for ABA-triggered chromatin opening in guard cells and implicate the inhibition of a set of bHLH-type transcription factors in controlling ABA-repressed chromatin. Moreover, we demonstrate that ABA and CO2 induce distinct programs of chromatin remodeling. We provide insight into the control of guard cell chromatin dynamics and propose that ABA-induced chromatin remodeling primes the genome for abiotic stress resistance. Overall design: We used the Assay for Transposaes Accessible Chromatin with sequencing (ATAC-seq) to profile genome-wide patterns of chromatin accessibility in different Arabidopsis thaliana cell-types (seedling roots, mesophyll cells, and guard cells), and following exposure of plants to different treatments (abscisic acid, 100 ppm atmospheric carbon dioxide, or 1000 ppm atmospheric carbon dioxide). Nuclei from different cell-types were purified using FACS performed on samples derived form transgenic plants expressing cell-type specific nuclear fluorescent markers (Histone H2B tagged with EGFP). All experiments were performed with a minimum of 2 biological replicates per cell-type and treatment.