BES1 and BZR1 Redundantly Promote Phloem and Xylem Differentiation.

Vascular development is a good model for studying cell differentiation in plants. Two conductive tissues, the xylem and phloem, are derived from common stem cells known as procambial/cambial cells. Glycogen synthase kinase 3 proteins (GSK3s) play crucial roles in maintaining procambial/cambial cells by suppressing their differentiation into xylem or phloem cells. We previously designed an in vitro culture system for analyzing vascular cell differentiation named VISUAL (Vascular cell Induction culture System Using Arabidopsis Leaves). Using this system, we found that the transcription factor BRI1-EMS-SUPPRESSOR 1 (BES1) functions as a downstream target of GSK3s during xylem differentiation. However, the function of BES1 in vascular development remains largely unknown. Here, we found that, in addition to xylem differentiation, BES1 positively regulates phloem differentiation downstream of GSK3s. Transcriptome analysis using VISUAL confirmed that BES1 promotes bi-directional differentiation of procambial cells into xylem and phloem cells. Genetic analysis of loss-of-function mutants newly generated using the CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9) system revealed that BRASSINAZOLE RESISTANT 1 (BZR1), the closest homolog of BES1, functions in vascular development redundantly with BES1. Notably, BZR1 has a weaker impact on vascular cell differentiation than BES1, suggesting that they contribute differentially to this process. In conclusion, our findings indicate that BES1 and BZR1 are key regulators of both xylem and phloem cell differentiation from vascular stem cells.