Brassinosteroids (BRs) are steroid hormones that are essential for plant growth and development. These hormones control the division, elongation and differentiation of various cell types throughout the entire plant life cycle. Our current understanding of the BR signaling pathway has mostly been obtained from studies using Arabidopsis thaliana as a model. In this context, the membrane steroid receptor BRI1 (BRASSINOSTEROID INSENSITIVE 1) binds directly to the BR ligand, triggering a signal cascade in the cytoplasm that leads to the transcription of BR-responsive genes that drive cellular growth. However, recent studies of the primary root have revealed distinct BR signaling pathways in different cell types and have highlighted cell-specific roles for BR signaling in controlling adaptation to stress. In this Review, we summarize our current knowledge of the spatiotemporal control of BR action in plant growth and development, focusing on BR functions in primary root development and growth, in stem cell self-renewal and death, and in plant adaption to environmental stress.

Understanding stem cell regulatory circuits is the next challenge in plant biology, as these cells are essential for tissue growth and organ regeneration in response to stress. In the Arabidopsis primary root apex, stem cell-specific transcription factors BRAVO and WOX5 co-localize in the quiescent centre (QC) cells, where they commonly repress cell division so that these cells can act as a reservoir to replenish surrounding stem cells, yet their molecular connection remains unknown. Genetic and biochemical analysis indicates that BRAVO and WOX5 form a transcription factor complex that modulates gene expression in the QC cells to preserve overall root growth and architecture. Furthermore, by using mathematical modelling we establish that BRAVO uses the WOX5/BRAVO complex to promote WOX5 activity in the stem cells. Our results unveil the importance of transcriptional regulatory circuits in plant stem cell development.

SUMMARYRoot growth and development are essential features for plant survival and the preservation of terrestrial ecosystems. In the Arabidopsis primary root apex, stem-cell specific transcription factors BRAVO and WOX5 co-localize at the Quiescent Center (QC) cells, where they repress cell division so that these cells can act as a reservoir to replenish surrounding stem cells, yet their molecular connection remains unknown. Here, by using empirical evidence and mathematical modeling, we establish the precise regulatory and molecular interactions between BRAVO and WOX5. We found that BRAVO and WOX5 regulate each other besides forming a transcription factor complex in the QC necessary to preserve overall root growth and architecture. Our results unveil the importance of transcriptional regulatory circuits at the quiescent and stem cells to the control of organ initiation and growth of plant tissues.

HighlightBrassinosteroid-regulated transcription factor BES1 targets the BRL3 receptor gene and finely modulates its transcription in the vascular and stem cells of the Arabidopsis primary root.

Summary Root analysis is essential for both academic and agricultural research. Despite the great advances in root phenotyping and imaging, calculating root length is still performed manually and involves considerable amounts of labor and time. To overcome these limitations, we developed MyROOT, a software for the semiautomatic quantification of root growth of seedlings growing directly on agar plates. Our method automatically determines the scale from the image of the plate, and subsequently measures the root length of the individual plants. To this aim, MyROOT combines a bottom‐up root tracking approach with a hypocotyl detection algorithm. At the same time as providing accurate root measurements, MyROOT also significantly minimizes the user intervention required during the process. Using Arabidopsis, we tested MyROOT with seedlings from different growth stages and experimental conditions. When comparing the data obtained from this software with that of manual root measurements, we found a high correlation between both methods (R2 = 0.997). When compared with previous developed software with similar features (BRAT and EZ‐Rhizo), MyROOT offered an improved accuracy for root length measurements. Therefore, MyROOT will be of great use to the plant science community by permitting high‐throughput root length measurements while saving both labor and time.

28Root analysis is essential for both academic and agricultural research. Despite the great 29 advances in root phenotyping and imaging however, calculating root length is still 30 performed manually and involves considerable amounts of labor and time. To overcome 31 these limitations, we have developed MyROOT, a novel software for the semi-automatic 32 quantification of root growth of seedlings growing directly in agar plates. Our method 33 automatically determines the scale from the image of the plate, and subsequently 34 measures the root length of the individual plants. To this aim, MyROOT combines a 35 bottom-up root tracking approach with a hypocotyl detection algorithm. At the same time 36as providing accurate root measurements, MyROOT also significantly minimizes the user 37 intervention required during the process. Using Arabidopsis, we tested MyROOT with 38 seedlings from different growth stages. Upon comparing the data obtained using this 39 software with that of manual root measurements, we found that there are no significant 40 differences (t-test, p-value < 0.05). Thus, MyROOT will be of great aid to the plant 41 science community by permitting high-throughput root length measurements while 42 saving on both labor and time. 43 44 48 impact of roots in agriculture 1 . As such, generating tools for precise, high-throughput 49 phenotyping and imaging of the root is essential for plant research and agriculture. Even 50 phenotyping facilities such as the ones available in the European Plant Phenotypic 51 Network (http://www.plant-phenotyping-network.eu/) have started to implement tools for 52 the massive screening of roots.53Roots provide the necessary structural and functional support for the incorporation of 54 nutrients and water from the soil. In Arabidopsis thaliana (Arabidopsis), the primary root 55 has a very simplified anatomy that makes it very amenable for genetic and microscopic 56 analyses 2-4 . Different root cell lineages are derived from the activity of a group of stem 57 cells located at the root apex. Here, the stem cell niche is formed by a few (3-7) quiescent 58 center (QC) cells that occasionally divide asymmetrically to renew themselves and to 59 3 form daughter stem cells. From the root apex, these cells actively divide in the 60 meristematic zone, and before exiting the cell cycle in the transition zone, continue to 61 elongate and differentiate in spatially separated regions of the root. In this way, primary 62 root growth is determined by the balance between cell division and cell elongation within 63 the different zones of the root 5-8 . 64The most straightforward symptom of abnormal root growth or development can be 65 identified by examining the length of the primary root in seedlings. Abnormalities in 66 length can usually be observed and measured just five to six days after germination 67 (DAG), where still reflect their embryonic origin 9 . Growth defects in the primary root of 68 seedlings are not only consistent with overall growth defects, but also persistent along the 6...

In animals and plants, stem-cell niches are local microenvironments that are tightly regulated to preserve their unique identity while communicating with adjacent cells that will give rise to specialized cell types. In the primary root of Arabidopsis thaliana, two transcription factors, BRAVO and WOX5, among others, are expressed in the stem-cell niche. Intriguingly, BRAVO, a repressor of quiescent center divisions, confines its own gene expression to the stem-cell niche, as evidenced in a bravo mutant background. Here, we propose through mathematical modeling that BRAVO confines its own expression domain to the stem-cell niche by attenuating a WOX5-dependent diffusible activator of BRAVO. This negative feedback drives WOX5 activity to be spatially restricted as well. The results show that WOX5 diffusion and sequestration by binding to BRAVO are sufficient to drive the experimentally observed confined BRAVO expression at the stem-cell niche. We propose that the attenuation of a diffusible activator can be a general mechanism acting at other stem-cell niches to spatially confine genetic activity to a small region while maintaining signaling within them and with the surrounding cells.

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