Auxin requirements for a meristematic state in roots depend on a dual brassinosteroid function

Root meristem organization is maintained by an interplay between hormone signaling pathways that both interpret and determine their accumulation and distribution. The interacting hormones Brassinosteroids (BR) and auxin control the number of meristematic cells in the Arabidopsis root. BR was reported both to promote auxin signaling input and to repress auxin signaling output. Whether these contradicting molecular outcomes co-occur and what their significance in meristem function is remain unclear. Here, we established a dual effect of BR on auxin, with BR simultaneously promoting auxin biosynthesis and repressing auxin transcriptional output, which is essential for meristem maintenance. Blocking BR-induced auxin synthesis resulted in rapid BR-mediated meristem loss. Conversely, plants with reduced BR levels were resistant to a critical loss of auxin biosynthesis, maintaining their meristem morphology. In agreement, injured root meristems, which rely solely on local auxin synthesis, regenerated when both auxin and BR synthesis were inhibited. Use of BIN2 as a tool to selectively inhibit BR signaling yielded meristems with distinct phenotypes depending on the perturbed tissue: meristem reminiscent either of BR-deficient mutants or of high BR exposure. This enabled mapping of the BR-auxin interaction that maintains the meristem to the outer epidermis and lateral root cap tissues and demonstrated the essentiality of BR signaling in these tissues for meristem response to BR. BR activity in internal tissues however, proved necessary to control BR levels. Together, we demonstrate a basis for inter-tissue coordination and how a critical ratio between these hormones determines the meristematic state. Arabidopsis (Arabidopsis thaliana) seedlings in the Columbia-0 (Col-0) background, were ‎germinated on one-half-strength Murashige and Skoog medium supplemented with 0.2% ‎‎(w/v) sucrose for 3 days. 3-day-old seedlings were transferred to treatments (mock, 0.8nM ‎BL, 4 µM Kyn and their combination, and harvested after 12 h, 24 h and 48 h.Total RNA was ‎extracted from roots. RNA sequencing and analysis were performed at the Technion ‎Genome Center (LSE). 24 samples (2 biological replicates) were prepared using the CEL-‎Seq2 sample preparation protocol (Hashimshony, et al., 2016), and sequenced on a paired ‎end run Illumina HiSeq 2500, to obtain 15 bp read 1 and 50 bp read 2. ‎