Assembling and remodelling the cell wall is essential for plant development. Cell wall dynamics are controlled by cell wall proteins, polysaccharide biosynthesis, and a variety of sensor and receptor systems. LecRK-I.9, an Arabidopsis thaliana plasma membrane-localized lectin receptor kinase, was previously shown to be involved in cell wall-plasma membrane contacts and to play roles in plant-pathogen interactions, but until now its role in development was not known. LecRK-I.9 is transcribed at a high level in root tissues including the pericycle. Comparative transcript profiling of a loss-of-function mutant versus the wild type identified LecRK-I.9 as a regulator of cell wall metabolism. Consistently, lecrk-I.9 mutants displayed an increased pectin methylesterification level correlated with decreased pectin methylesterase and increased polygalacturonase activities. Also, LecRK-I.9 negatively impacted lateral root development through the direct or indirect regulation of genes encoding (i) cell wall remodelling proteins during early events of lateral root initiation, and (ii) cell wall signalling peptides (CLE2 and CLE4) repressing lateral root emergence and growth. Furthermore, low nitrate reduced LecRK-I.9 expression in roots, particularly in the lateral root emergence zone: even in these conditions, the control of CLE2 and CLE4 expression is maintained. Altogether, the results show that LecRK-I.9 is a key player in negatively regulating both pre-branch site formation and lateral root emergence.

In Arabidopsis thaliana, lateral root (LR) development requires spatial accommodation responses in overlying endodermal cells. This includes loss of cell volume whilst maintaining membrane integrity to allow the expansion of the underlying LR primordia (LRPs). These accommodation responses are regulated by auxin-mediated signaling, specifically through Aux/IAA proteins, involving IAA3/SHY2. Plants that express a stabilized version of SHY2 (shy2-2) in differentiated endodermal cells, CASP1pro::shy2-2 plants, fail to make LRs. Exogenous treatment with 1-naphthaleneacetic acid (NAA) was reported to partially restore LR formation in this spatial accommodation mutant. Using treatments with auxins having different transport properties, such as NAA, indole-3-acetic acid (IAA), and 2,4-dichlorophenoxyacetic acid (2,4-D), we assessed the ability of each auxin to rescue LR formation in CASP1pro::shy2-2 roots. This revealed that IAA is the most effective in partially restoring LR development, NAA is effective in inducing LRPs but cannot maintain their canonical phenotype, whereas 2,4-D induces non-controlled cell divisions. In addition, we show that in CASP1pro::shy2-2 roots, AUX1 appears to be repressed in the zone where oscillation of the auxin response has been described. Our study advances the understanding of auxin-regulated spatial accommodation mechanisms during LRP formation and highlights the complex interplay of auxin transport and signaling in bypassing the endodermal constraints.

AbstractReactive oxygen species (ROS) function as key signals in plants to enable adaptation to environmental stresses. Plant roots respond to transient water stress by temporarily ceasing branching using the acclimative response xerobranching1. In this study, we report that a rapid ROS burst regulates Xerobranching by inducing multimerization of auxin repressor protein IAA3/SHY2. Mutations in specific cysteine residues in IAA3/SHY2 disrupt redox-mediated multimerization and interaction with co-repressor TPL, but not with auxin response partner ARF7 and auxin receptor TIR1. ROS-mediated oligomerization of IAA3/SHY2 is required for efficient ARF mediated target gene repression during Xerobranching and lateral root emergence. We demonstrate that AUX/IAA proteins vary in their redox mediated multimerization, revealing a new auxin response regulatory mechanism that directly connects ROS sensing to auxin signalling. Our study reveals how ROS, auxin and water stress intersect to shape acclimative responses in plant roots and maintain their phenotypic plasticity.

The root system of plants is a vital part for successful development and adaptation to different soil types and environments. A major determinant of the shape of a plant root system is the formation of lateral roots, allowing for expansion of the root system. Arabidopsis thaliana, with its simple root anatomy, has been extensively studied to reveal the genetic program underlying root branching. However, to get a more general understanding of lateral root development, comparative studies in species with a more complex root anatomy are required. Here, by combining optimized clearing methods and histology, we describe an atlas of lateral root development in Brachypodium distachyon, a wild, temperate grass species. We show that lateral roots initiate from enlarged phloem pole pericycle cells and that the overlying endodermis reactivates its cell cycle and eventually forms the root cap. In addition, auxin signaling reported by the DR5 reporter was not detected in the phloem pole pericycle cells or young primordia. In contrast, auxin signaling was activated in the overlying cortical cell layers, including the exodermis. Thus, Brachypodium is a valuable model to investigate how signaling pathways and cellular responses have been repurposed to facilitate lateral root organogenesis.

The development of lateral roots starts with a round of anticlinal, asymmetric cell divisions in lateral root founder cells in the pericycle, deep within the root. The reorientation of the cell division plane occurs in parallel with changes in cell shape and needs to be coordinated with its direct neighbor, the endodermis. This accommodation response requires the integration of biochemical and mechanical signals in both cell types. Recently, it was reported that dynamic changes in the cytoskeleton and possibly the cell wall are part of the molecular mechanism required to correctly orient and position the cell division plane. Here we discuss the latest progress made towards our understanding of the regulation of cell shape and division plane orientation underlying lateral root initiation in Arabidopsis.

Abstract The underground development of the plant root system is a complex and plastic process. It enables the plants to mine hydromineral resources while dealing with changes in their environment. Root branching via lateral root formation is considered as an important trait because it greatly influences root system architecture and impacts plant growth and crop performance. In the past few years, major progresses in understanding the mechanisms of root branching have been made. Technical advances lift the veil on multiple molecular and cellular processes that contribute to the robust formation of new lateral roots in plant models and to its response to endogenous and environmental cues. Developing translational approaches should increase in the coming decades to allow a global and integrative vision of root system development and the mechanisms of root branching in various plant species. Moreover, exploring the genetic diversity that modulates this fundamental process opens the way to breeding crops with more efficient root system architectures. Key Concepts Lateral root formation significantly influences root system architecture, a key trait for plant interaction with the soil and especially, plant nutrition. Lateral root primordium initiation, development and emergence from the primary root have been well described in several plant model species. Lateral root formation is highly regulated. Many molecular mechanisms involved in this organogenesis process are known in detail, especially in the plant model Arabidopsis thaliana . Among these, the phytohormone auxin is a key regulatory signal influencing each step of lateral root formation. A complex regulatory network, integrating intercellular signals and mechanical constraints, triggers regularly spaced initiation of lateral root formation, allows robust organ patterning, and controls the lateral root primordium growth through the overlaying primary root tissues. Endogenous and environmental signals modulate lateral root formation kinetics and emergence, yielding plasticity in root system development. Comparison of lateral root development in different plant species reveals specific and conserved pathways. Genetic diversity can be explored and used to breed crops with optimised root branching properties.

Abstract Lateral root organogenesis is a key process in the development of a plant’s root system and its adaptation to the environment. During lateral root formation, an early phase of cell proliferation first produces a four-cell-layered primordium, and only from this stage onwards is a root meristem-like structure, expressing root stem cell niche marker genes, being established in the developing organ. Previous studies reported that the gene regulatory network controlling lateral root formation is organized into two subnetworks whose mutual inhibition may contribute to organ patterning. PUCHI encodes an AP2/ERF transcription factor expressed early during lateral root primordium development and required for correct lateral root formation. To dissect the molecular events occurring during this early phase, we generated time-series transcriptomic datasets profiling lateral root development in puchi-1 mutants and wild types. Transcriptomic and reporter analyses revealed that meristem-related genes were expressed ectopically at early stages of lateral root formation in puchi-1 mutants. We conclude that, consistent with the inhibition of genetic modules contributing to lateral root development, PUCHI represses ectopic establishment of meristematic cell identities at early stages of organ development. These findings shed light on gene network properties that orchestrate correct timing and patterning during lateral root formation.

An efficient purification of recombinant proteins often requires a high ratio of recombinant to host proteins. In plants, Ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) is the most abundant leaf protein, thus strongly impacting purification yield. Here, we describe a simple and robust purification procedure for recombinant proteins based on a differential precipitation of RuBisCO. In this context, four Legume lectin domains of Arabidopsis thaliana which belong to receptor-like kinases and cell wall proteins were produced from Nicotiana benthamiana leaves. The recombinant proteins exhibit a unique lectin domain consisting of around 250 amino acid residues with several predicted N-glycosylation sites and a six His-tag at the N-terminus. After ammonium sulphate precipitation of total soluble proteins, depletion of RuBisCO was obtained using citrate and succinate buffers during the salting-in step: this depletion was pH-dependent and the presence of di- or tri-carboxylic acids was required. The depleted protein extracts were then subjected to two chromatographic steps which were used in the negative mode to submit a protein fraction enriched as much as possible in recombinant lectin domains to a third chromatographic step (immobilized metal-ion chromatography). Three of the Legume lectin domains were purified near to homogeneity and revealed multiple N-glycosylation isoforms, particularly those from receptor-like kinases, which were characterised using specific lectins and deglycosylation enzymes. The production and purification of recombinant lectin domains will facilitate their biochemical characterisation in the context of cell-to-cell signalling and cell wall organisation.

Lateral roots (LRs) are crucial for increasing the surface area of root systems to explore heterogeneous soil environments. Major advances have recently been made in the model plant arabidopsis (Arabidopsis thaliana) to elucidate the cellular basis of LR development and the underlying gene regulatory networks (GRNs) that control the morphogenesis of the new root organ. This has provided a foundation for understanding the sophisticated adaptive mechanisms that regulate how plants pattern their root branching to match the spatial availability of resources such as water and nutrients in their external environment. We review new insights into the molecular, cellular, and environmental regulation of LR development in arabidopsis.

La paroi donne une enveloppe rigide a la cellule vegetale. Elle renferme des polysaccharides (cellulose, hemicelluloses, pectines), des lignines dans certains types cellulaires, des proteines structurales et enzymatiques. La paroi est une structure dynamique: les parois sont modifiees au cours du developpement et en reponse aux contraintes environnementales. Les proteines sont des acteurs de cette dynamique en participant a l'assemblage et au remodelage des polysaccharides, a la signalisation cellulaire. La paroi permet notamment aux cellules de resister a la pression de turgescence, force motrice de l'elongation cellulaire: les etapes de relaxation, d'integration de nouveaux constituants et de consolidation de la paroi doivent etre parfaitement coordonnees pour maintenir les structures parietales. Un systeme de surveillance et des senseurs du statut de la paroi pourraient assurer cette coordination. Nous sommes interesses par un recepteur lectine-kinase (LecRK-I.9) d'Arabidopsis thaliana dont le domaine extracellulaire est de type lectine de Legumineuses. Il est un acteur potentiel d'un systeme de surveillance du statut de la paroi. Les questions posees dans ce travail sont: (i) dans quels processus de developpement LecRK-I.9 peut-il etre implique ? (ii) quelles sont les regulations que ciblent LecRK-I.9 ? (iii) quelle est la nature des ligands de LecRK-I.9 ? L'expression de LecRK-I.9 est preponderante dans les tissus racinaires et nous avons pu montrer que LecRK-I.9 est implique dans les processus d'initiation et d'emergence des racines laterales et adventives pour lesquels des remodelages parietaux sont necessaires. Il apparait comme regulateur negatif de ces processus. En effet, chez les plantes lecrk-I.9, l'expression de genes codant des peptides parietaux CEP, regulateurs precoces de l'initiation des racines laterales, est deregulee, de meme pour de nombreuses enzymes travaillant de facon concertee a un relâchement des structures parietales. Les plantes lecrk-I.9 presentent ainsi des parois modifiees dans leur composition en polysaccharides. Un dommage cause a la paroi a ete obtenu par un traitement inhibiteur de la biosynthese de cellulose. En particulier, des depots de lignine ectopique se mettent en place dans les apex racinaires sous le controle de l'acide jasmonique (JA) et des especes reactives de l'oxygene (ROS). Nous avons pu montrer que LecRK-I.9 controle les teneurs en JA dans ce processus. De plus, via la regulation JA, LecRK-I.9 controle l'expression de genes codant des proteines et des peptides parietaux, mais egalement des proteines de detoxification des ROS. L'ensemble des resultats suggerent que LecRK-I.9 est un regulateur de la dynamique parietale avec pour cibles, les teneurs en JA, l'homeostasie des ROS, les enzymes de remodelage des polysaccharides. Le travail s'oriente maintenant vers les consequences de modifications de la composition parietale sur la nutrition minerale en fer. En effet, les plantes lecrk-I.9 montrent une forte accumulation de fer parietal. Enfin, LecRK-I.9 est associe aux brins d'Hecht, en particulier dans les points d'ancrage parietaux. L'interaction entre domaines lectine et composes parietaux a l'aide de puces a oligosaccharides pourrait determiner la participation de LecRK-I.9 a une continuite structurale entre paroi et plasmalemme.

Lectins are fundamental to plant life and have important roles in cell-to-cell communication; development and defence strategies. At the cell surface; lectins are present both as soluble proteins (LecPs) and as chimeric proteins: lectins are then the extracellular domains of receptor-like kinases (LecRLKs) and receptor-like proteins (LecRLPs). In this review; we first describe the domain architectures of proteins harbouring G-type; L-type; LysM and malectin carbohydrate-binding domains. We then focus on the functions of LecPs; LecRLKs and LecRLPs referring to the biological processes they are involved in and to the ligands they recognize. Together; LecPs; LecRLKs and LecRLPs constitute versatile recognition systems at the cell surface contributing to the detection of symbionts and pathogens; and/or involved in monitoring of the cell wall structure and cell growth.

SUMMARYIn cereals, the root system is mainly composed of post‐embryonic shoot‐borne roots, named crown roots. The CROWN ROOTLESS1 (CRL1) transcription factor, belonging to the ASYMMETRIC LEAVES2‐LIKE/LATERAL ORGAN BOUNDARIES DOMAIN (ASL/LBD) family, is a key regulator of crown root initiation in rice (Oryza sativa). Here, we show that CRL1 can bind, both in vitro and in vivo, not only the LBD‐box, a DNA sequence recognized by several ASL/LBD transcription factors, but also another not previously identified DNA motif that was named CRL1‐box. Using rice protoplast transient transactivation assays and a set of previously identified CRL1‐regulated genes, we confirm that CRL1 transactivates these genes if they possess at least a CRL1‐box or an LBD‐box in their promoters. In planta, ChIP‐qPCR experiments targeting two of these genes that include both a CRL1‐ and an LBD‐box in their promoter show that CRL1 binds preferentially to the LBD‐box in these promoter contexts. CRISPR/Cas9‐targeted mutation of these two CRL1‐regulated genes, which encode a plant Rho GTPase (OsROP) and a basic helix–loop–helix transcription factor (OsbHLH044), show that both promote crown root development. Finally, we show that OsbHLH044 represses a regulatory module, uncovering how CRL1 regulates specific processes during crown root formation.