Unlike activation of target genes in response to abscisic acid (ABA), how MYB96 transcription factor represses ABA-repressible genes to further enhance ABA responses remains unknown. Here, we show MYB96 interacts with the histone modifier HDA15 to suppress negative regulators of early ABA signaling. The MYB96-HDA15 complex co-binds to the promoters of a subset ofRHO GTPASE OF PLANTS(ROP) genes,ROP6,ROP10, andROP11, and represses their expression by removing acetyl groups of histone H3 and H4 from the cognate regions, particularly in the presence of ABA. In support,HDA15-deficient mutants display reduced ABA sensitivity and are susceptible to drought stress with derepression of theROPgenes, as observed in themyb96-1mutant. Biochemical and genetic analyses show that MYB96 and HDA15 are interdependent in the regulation ofROPsuppression. Thus, MYB96 confers maximal ABA sensitivity by regulating both positive and negative regulators of ABA signaling through distinctive molecular mechanisms.

Background: Plants have a remarkable reprogramming potential, which facilitates plant regeneration, especially from a single cell. Protoplasts have the ability to form a cell wall and undergo cell division, allowing whole plant regeneration. With the growing need for protoplast regeneration in genetic engineering and genome editing, fundamental studies that enhance our understanding of cell cycle re-entry, pluripotency acquisition, and de novo tissue regeneration are essential. To conduct these studies, a reproducible and efficient protoplast regeneration method using model plants is necessary. Results: Here, we optimized cell and tissue culture methods for improving protoplast regeneration efficiency in Arabidopsis thaliana. Protoplasts were isolated from whole seedlings of four different Arabidopsis ecotypes including Columbia (Col-0), Wassilewskija (Ws-2), Nossen (No-0), and HR (HR-10). Among these ecotypes, Ws-2 showed the highest potential for protoplast regeneration. A modified thin alginate layer was applied to the protoplast culture at an optimal density of 1 × 10⁶ protoplasts/mL. Following callus formation and de novo shoot regeneration, the regenerated inflorescence stems were used for de novo root organogenesis. The entire protoplast regeneration process was completed within 15 weeks. The in vitro regenerated plants were fertile and produced morphologically normal progenies. Conclusion: The cell and tissue culture system optimized in this study for protoplast regeneration is efficient and reproducible. This method of Arabidopsis protoplast regeneration can be used for fundamental studies on pluripotency establishment and de novo tissue regeneration.

The histone lysine methyltransferase ATXR2 promotes cellular dedifferentiation in Arabidopsis thaliana .