While apical growth in plants initiates upon seed germination, radial growth is only primed during early ontogenesis in procambium cells and activated later by the vascular cambium 1 . Although it is not known how radial growth is organized and regulated in plants, this system resembles the developmental competence observed in some animal systems, in which pre-existing patterns of developmental potential are established early on 2,3 . Here we show that the initiation of radial growth occurs around early protophloem sieve element (PSE) cell files of the root procambial tissue in Arabidopsis. In this domain cytokinin signalling promotes expression of a pair of novel mobile transcription factors, PHLOEM EARLY DOF (PEAR1, PEAR2) and their four homologs (DOF6, TMO6, OBP2 and HCA2), collectively called PEAR proteins. The PEAR proteins form a short-range concentration gradient peaking at PSE and activating gene expression that promotes radial growth. The expression and function of PEAR proteins are antagonized by well-established polarity transcription factors, HD-ZIP III 4 , whose expression is concentrated in the more internal domain of radially non-dividing procambial cells by the function of auxin and mobile miR165/166. The PEAR proteins locally promote transcription of their inhibitory HD-ZIP III genes, thereby establishing a negative feedback loop that forms a robust boundary demarking the zone of cell divisions. Taken together, we have established a network, in which the PEAR -HD-ZIP III module integrates spatial information of the hormonal domains and miRNA gradients during root procambial development, to provide adjacent zones of dividing and more quiescent cells as a foundation for further radial growth. Cambial growth in plants is initiated within the procambial tissues of the apical meristems through periclinal (i.e. longitudinal) divisions associated with formation of the vascular tissues xylem and phloem 1 (Extended Data Fig. 1a). It has been established that during procambial development in Arabidopsis roots there are distinct domains for high auxin and cytokinin signalling, which mark the regions for further development of xylem and phloem/procambium, respectively 5-8 . To accurately map the spatial distribution of the periclinal divisions, we established a new nomenclature for the root procambial cells, including PSE-lateral neighbours (PSE-LN) as cells directly contacting both PSE and the pericycle, the outer procambial cells (OPC) as procambial cells adjacent to the pericycle but not contacting PSE, and SE-internal neighbours (PSE-IN) as cells located internal to and directly contacting PSE (Fig. 1a). Both the PSE cell and PSE-LN showed higher activity of periclinal cell division than the OPC and PSE-IN (Fig. 1b, Extended Data Fig. 1b-d and Supplementary Information).We observed virtually no periclinal divisions in metaxylem (MX) and internal procambial cells (IPC) (Fig. 1b). Furthermore, blocking symplastic transport genetically 9 between the PSE and the surrounding cells results in a dramatic reduct...

Lateral root formation is an important determinant of root system architecture. In Arabidopsis, lateral roots originate from pericycle cells, which undergo a program of morphogenesis to generate a new lateral root meristem. Despite its importance for root meristem organization, the onset of quiescent center (QC) formation during lateral root morphogenesis remains unclear. Here, we used live 3D confocal imaging to monitor cell organization and identity acquisition during lateral root development. Our dynamic observations revealed an early morphogenesis phase and a late meristem formation phase as proposed in the bi-phasic growth model. Establishment of lateral root QCs coincided with this developmental phase transition. QC precursor cells originated from the outer layer of stage II lateral root primordia, within which the SCARECROW (SCR) transcription factor was specifically expressed. Disrupting SCR function abolished periclinal divisions in this lateral root primordia cell layer and perturbed the formation of QC precursor cells. We conclude that de novo QC establishment in lateral root primordia operates via SCR-mediated formative cell division and coincides with the developmental phase transition.

Highlights d Lateral root founder cells (LRFCs) express TOLS2 peptide, inhibiting nearby LRFCs d The receptor RLK7 and transcription factor PUCHI make up the TOLS2 signaling pathway d DR5, an LRFC marker, is occasionally activated in proximity to pre-existing LRFCs d TOLS2-RLK7-PUCHI inhibits LRFC identity in proximal cells, ensuring proper LR spacing SUMMARYIn plants, the position of lateral roots (LRs) depends on initiation sites induced by auxin. The domain of high auxin response responsible for LR initiation stretches over several cells, but only a pair of pericycle cells (LR founder cells) will develop into LRs.In this work, we identified a signaling cascade controlling LR formation through lateral inhibition. It comprises a peptide hormone TARGET OF LBD SIXTEEN 2 (TOLS2), its receptor RLK7, and a downstream transcription factor PUCHI. TOLS2 is expressed at the LR founder cells and inhibits LR initiation. Time-lapse imaging of auxin-responsive DR5:LUCIFERASE reporter expression revealed that occasionally two pairs of LR founder cells are specified in close proximity even in wild-type and that one of them exists only transiently and disappears in an RLK7-dependent manner. We propose that the selection of LR founder cells by the peptide hormone-receptor cascade ensures proper LR spacing.