Formative Cell Divisions Research Articles

ARGONAUTE10 controls cell fate specification and formative cell divisions in the Arabidopsis root

A key question in plant biology is how oriented cell divisions are integrated with patterning mechanisms to generate organs with adequate cell type allocation. In the root vasculature, a gradient of miRNA165/6 controls the abundance of HD-ZIP III transcription factors, which in turn control cell fate and spatially restrict vascular cell proliferation to specific cells. Here, we show that vascular development requires the presence of ARGONAUTE10, which is thought to sequester miRNA165/6 and protect HD-ZIP III transcripts from degradation. Our results suggest that the miR165/6-AGO10-HDZIP III module acts by buffering cytokinin responses and restricting xylem differentiation. Mutants of AGO10 show faster growth rates and strongly enhanced survival under severe drought conditions. However, this superior performance is offset by markedly increased variation and phenotypic plasticity in sub-optimal carbon supply conditions. Thus, AGO10 is required for the control of formative cell division and coordination of robust cell fate specification of the vasculature, while altering its expression provides a means to adjust phenotypic plasticity.

SHR and SCR coordinate root patterning and growth early in the cell cycle

Precise control of cell division is essential for proper patterning and growth during the development of multicellular organisms. Coordination of formative divisions that generate new tissue patterns with proliferative divisions that promote growth is poorly understood. SHORTROOT (SHR) and SCARECROW (SCR) are transcription factors that are required for formative divisions in the stem cell niche of Arabidopsis roots1,2. Here we show that levels of SHR and SCR early in the cell cycle determine the orientation of the division plane, resulting in either formative or proliferative cell division. We used 4D quantitative, long-term and frequent (every 15 min for up to 48 h) light sheet and confocal microscopy to probe the dynamics of SHR and SCR in tandem within single cells of living roots. Directly controlling their dynamics with an SHR induction system enabled us to challenge an existing bistable model3 of the SHR–SCR gene-regulatory network and to identify key features that are essential for rescue of formative divisions in shr mutants. SHR and SCR kinetics do not align with the expected behaviour of a bistable system, and only low transient levels, present early in the cell cycle, are required for formative divisions. These results reveal an uncharacterized mechanism by which developmental regulators directly coordinate patterning and growth.

GRAS transcription factors regulate cell division planes in moss overriding the default rule

Plant cells are surrounded by a cell wall and do not migrate, which makes the regulation of cell division orientation crucial for development. Regulatory mechanisms controlling cell division orientation may have contributed to the evolution of body organization in land plants. The GRAS family of transcription factors was transferred horizontally from soil bacteria to an algal common ancestor of land plants. SHORTROOT (SHR) and SCARECROW (SCR) genes in this family regulate formative periclinal cell divisions in the roots of flowering plants, but their roles in nonflowering plants and their evolution have not been studied in relation to body organization. Here, we show that SHR cell autonomously inhibits formative periclinal cell divisions indispensable for leaf vein formation in the moss Physcomitrium patens, and SHR expression is positively and negatively regulated by SCR and the GRAS member LATERAL SUPPRESSOR, respectively. While precursor cells of a leaf vein lacking SHR usually follow the geometry rule of dividing along the division plane with the minimum surface area, SHR overrides this rule and forces cells to divide nonpericlinally. Together, these results imply that these bacterially derived GRAS transcription factors were involved in the establishment of the genetic regulatory networks modulating cell division orientation in the common ancestor of land plants and were later adapted to function in flowering plant and moss lineages for their specific body organizations.

(Don't) Look Up!: Is short-root just a short-root plant?

SHORT-ROOT (SHR) is a mobile transcription factor that plays important roles in ground tissue patterning, stem cell niche specification and maintenance, and vascular development in Arabidopsis roots. Although mRNA and protein of SHR are also found in hypocotyls, inflorescence stems, and leaves, its role in the above-ground organs has been less explored. In most developmental cases, SHR, together with its partner SCARECROW (SCR), regulates the expression of downstream target genes in controlling formative and proliferative cell divisions. Accumulating evidence on the regulatory role of SHR in shoots suggests that SHR may also play key roles in the above-ground organs. Interestingly, recent work has provided new evidence that SHR is also required for cell elongation in the hypocotyl of the etiolated seedling. This suggests that the novel roles of SHR and SHR-mediated regulatory networks can be found in shoots. Furthermore, comparative research on SHR function in roots and shoots will broaden and deepen our understanding of plant growth and development.

Confocal Analysis of Arabidopsis Root Cell Divisions in 3D: A Focus on the Endodermis.

The development of multicellular organisms requires coordinated cell divisions for the production of diverse cell types and body plan elaboration and growth. There are two main types of cell divisions: proliferative or symmetric divisions, which produce more cells of a given type, and formative or asymmetric divisions, which produce cells of different types. Because plant cells are surrounded by cell walls, the orientation of plant cell divisions is particularly important in cell fate specification and tissue or organ morphology. The cellular organization of the Arabidopsis thaliana root makes an excellent tool to study how oriented cell division contributes to tissue patterning during organ development. To understand how division plane orientation in a specific genotype or growth condition may impact organ or tissue development, a detailed characterization of cell division orientation is required. Here we describe a confocal microscopy-based, live imaging method for Arabidopsis root tips to examine the 3D orientations of cell division planes and quantify formative, proliferative, and atypical endodermal cell divisions.

BAM1/2 receptor kinase signaling drives CLE peptide-mediated formative cell divisions in Arabidopsis roots

Cell division is often regulated by extracellular signaling networks to ensure correct patterning during development. In Arabidopsis, the SHORT-ROOT (SHR)/SCARECROW (SCR) transcription factor dimer activates CYCLIND6;1 (CYCD6;1) to drive formative divisions during root ground tissue development. Here, we show plasma-membrane-localized BARELY ANY MERISTEM1/2 (BAM1/2) family receptor kinases are required for SHR-dependent formative divisions and CYCD6;1 expression, but not SHR-dependent ground tissue specification. Root-enriched CLE ligands bind the BAM1 extracellular domain and are necessary and sufficient to activate SHR-mediated divisions and CYCD6;1 expression. Correspondingly, BAM-CLE signaling contributes to the restriction of formative divisions to the distal root region. Additionally, genetic analysis reveals that BAM-CLE and SHR converge to regulate additional cell divisions outside of the ground tissues. Our work identifies an extracellular signaling pathway regulating formative root divisions and provides a framework to explore this pathway in patterning and evolution.

The Cyclin CYCA3;4 Is a Postprophase Target of the APC/CCCS52A2 E3-Ligase Controlling Formative Cell Divisions in Arabidopsis.

The anaphase promoting complex/cyclosome (APC/C) controls unidirectional progression through the cell cycle by marking key cell cycle proteins for proteasomal turnover. Its activity is temporally regulated by the docking of different activating subunits, known in plants as CELL DIVISION PROTEIN20 (CDC20) and CELL CYCLE SWITCH52 (CCS52). Despite the importance of the APC/C during cell proliferation, the number of identified targets in the plant cell cycle is limited. Here, we used the growth and meristem phenotypes of Arabidopsis (Arabidopsis thaliana) CCS52A2-deficient plants in a suppressor mutagenesis screen to identify APC/CCCS52A2 substrates or regulators, resulting in the identification of a mutant cyclin CYCA3;4 allele. CYCA3;4 deficiency partially rescues the ccs52a2-1 phenotypes, whereas increased CYCA3;4 levels enhance the scored ccs52a2-1 phenotypes. Furthermore, whereas the CYCA3;4 protein is promptly broken down after prophase in wild-type plants, it remains present in later stages of mitosis in ccs52a2-1 mutant plants, marking it as a putative APC/CCCS52A2 substrate. Strikingly, increased CYCA3;4 levels result in aberrant root meristem and stomatal divisions, mimicking phenotypes of plants with reduced RETINOBLASTOMA-RELATED PROTEIN1 (RBR1) activity. Correspondingly, RBR1 hyperphosphorylation was observed in CYCA3;4 gain-of-function plants. Our data thus demonstrate that an inability to timely destroy CYCA3;4 contributes to disorganized formative divisions, possibly in part caused by the inactivation of RBR1.

Spatiotemporal Pattern of Ectopic Cell Divisions Contribute to Mis-Shaped Phenotype of Primary and Lateral Roots of katanin1 Mutant.

Pattern formation, cell proliferation, and directional cell growth, are driving factors of plant organ shape, size, and overall vegetative development. The establishment of vegetative morphogenesis strongly depends on spatiotemporal control and synchronization of formative and proliferative cell division patterns. In this context, the progression of cell division and the regulation of cell division plane orientation are defined by molecular mechanisms converging to the proper positioning and temporal reorganization of microtubule arrays such as the preprophase microtubule band, the mitotic spindle and the cytokinetic phragmoplast. By focusing on the tractable example of primary root development and lateral root emergence in Arabidopsis thaliana, genetic studies have highlighted the importance of mechanisms underlying microtubule reorganization in the establishment of the root system. In this regard, severe alterations of root growth, and development found in extensively studied katanin1 mutants of A. thaliana (fra2, lue1, and ktn1-2), were previously attributed to defective rearrangements of cortical microtubules and aberrant cell division plane reorientation. How KATANIN1-mediated microtubule severing contributes to tissue patterning and organ morphogenesis, ultimately leading to anisotropy in microtubule organization is a trending topic under vigorous investigation. Here we addressed this issue during root development, using advanced light-sheet fluorescence microscopy (LSFM) and long-term imaging of ktn1-2 mutant expressing the GFP-TUA6 microtubule marker. This method allowed spatial and temporal monitoring of cell division patterns in growing roots. Analysis of acquired multidimensional data sets revealed the occurrence of ectopic cell divisions in various tissues including the calyptrogen and the protoxylem of the main root, as well as in lateral root primordia. Notably the ktn1-2 mutant exhibited excessive longitudinal cell divisions (parallel to the root axis) at ectopic positions. This suggested that changes in the cell division pattern and the occurrence of ectopic cell divisions contributed significantly to pleiotropic root phenotypes of ktn1-2 mutant. LSFM provided evidence that KATANIN1 is required for the spatiotemporal control of cell divisions and establishment of tissue patterns in living A. thaliana roots.

Into another dimension: how streptophyte algae gained morphological complexity.

Land plants with elaborated three-dimensional (3D) body plans have evolved from streptophyte algae. The streptophyte algae are known to exhibit varying degrees of morphological complexity, ranging from single-celled flagellates to branched macrophytic forms exhibiting tissue-like organization. In this review, I discuss mechanisms by which, during evolution, filamentous algae may have gained 2D and eventually 3D body plans. There are, in principle, two mechanisms by which an additional dimension may be added to an existing algal filament or cell layer: first, by tip growth-mediated branching. An example of this mechanism is the emergence and polar expansion of root hairs from land plants. The second possibility is the rotation of the cell division plane. In this case, the plane of the forthcoming cell division is rotated within the parental cell wall. This type of mechanism corresponds to the formative cell division seen in meristems of land plants. This literature review shows that of the extant streptophyte algae, the Charophyceae and Coleochaetophyceae are capable of performing both mechanisms, while the Zygnematophyceae (the actual sister to land plants) show tip growth-based branching only. I finally discuss how apical cells with two or three cutting faces, as found in mosses, may have evolved from algal ancestors.

Overexpression of three related root-cap outermost-cell-specific C2H2-type zinc-finger protein genes suppresses the growth of Arabidopsis in an EAR-motif-dependent manner

The root meristem of Arabidopsis thaliana is protected by the root cap, the size of which is tightly regulated by the balance between the formative cell divisions and the dispersal of the outermost cells. We isolated an enhancer-tagged dominant mutant displaying the short and twisted root by the overexpression of ZINC-FINGER OF ARABIDOPSIS THALIANA1 (ZAT1) encoding an EAR motif-containing zinc-finger protein. The growth inhibition by ZAT1 was shared by ZAT4 and ZAT9, the ZAT1 homologues. The ZAT1 promoter was specifically active in the outermost cells of the root cap, in which ZAT1-GFP was localized when expressed by the ZAT1 promoter. The outermost cell-specific expression pattern of ZAT1 was not altered in the sombrero (smb) or smb bearskin1 (brn1) brn2 accumulating additional root-cap layers. In contrast, ZAT4-GFP and ZAT9- GFP fusion proteins were distributed to the inner root-cap cells in addition to the outermost cells where ZAT4 and ZAT9 promoters were active. Overexpression of ZAT1 induced the ectopic expression of PUTATIVE ASPARTIC PROTEASE3 involved in the programmed cell death. The EAR motif was essential for the growth inhibition by ZAT1. These results suggest that the three related ZATs might regulate the maturation of the outermost cells of the root cap.

The contribution of cell wall remodeling and signaling to lateral organs formation

Lateral organs are formed in plants by post embryonic developmental programs. Leaves, and flowers differentiate from the shoot apical meristem and lateral roots from the primary root pericycle meristem. Adventitious roots are roots formed from non-root lateral meristematic tissues, mostly the cambium, in many cases in response to stress signals. The ability of plants to regenerate adventitious roots is fundamental for selection and breading programs which are based on vegetative propagation of elite clones. Thus, recalcitrant plants, losing their rooting capability, may form a genuine commercial barrier in agricultural and forestry improvement programs. Some cellular mechanisms underlying adventitious root formation have been revealed, but much is yet to be clarified. The plant primary cell wall is a dynamic organ that can change its form, and perceive and relay molecular signals inward and outward during certain stages of development in particular cells. Therefore, before the secondary cell wall is deposited and plants become the wood from which walls and furniture are built, and the fibers from which cloths are woven, primary cell walls actively participate in plant cell differentiation and developmental programs. While auxin is a major regulator, cell walls are important in regulating coherent formative cell division and synchronized polar elongation of cell lineages that are necessary for lateral organ induction and formation, and collaborative cell functioning. Nevertheless, little is known of how cell wall changes are molecularly sensed and translated to intracellular signals during differentiation of adventitious roots. Here we summarize recent data linking, directly or indirectly, cell wall events to auxin signaling and to lateral or adventitious root induction and formation.

Stomatal Complex Development and F-Actin Organization in Maize Leaf Epidermis Depend on Cellulose Synthesis.

Cellulose microfibrils reinforce the cell wall for morphogenesis in plants. Herein, we provide evidence on a series of defects regarding stomatal complex development and F-actin organization in Zea mays leaf epidermis, due to inhibition of cellulose synthesis. Formative cell divisions of stomatal complex ontogenesis were delayed or inhibited, resulting in lack of subsidiary cells and frequently in unicellular stomata, with an atypical stomatal pore. Guard cells failed to acquire a dumbbell shape, becoming rounded, while subsidiary cells, whenever present, exhibited aberrant morphogenesis. F-actin organization was also affected, since the stomatal complex-specific arrays were scarcely observed. At late developmental stages, the overall F-actin network was diminished in all epidermal cells, although thick actin bundles persisted. Taken together, stomatal complex development strongly depends on cell wall mechanical properties. Moreover, F-actin organization exhibits a tight relationship with the cell wall.

PLETHORA transcription factors orchestrate de novo organ patterning during Arabidopsis lateral root outgrowth

Plant development is characterized by repeated initiation of meristems, regions of dividing cells that give rise to new organs. During lateral root (LR) formation, new LR meristems are specified to support the outgrowth of LRs along a new axis. The determination of the sequential events required to form this new growth axis has been hampered by redundant activities of key transcription factors. Here, we characterize the effects of three PLETHORA (PLT) transcription factors, PLT3, PLT5, and PLT7, during LR outgrowth. In plt3plt5plt7 triple mutants, the morphology of lateral root primordia (LRP), the auxin response gradient, and the expression of meristem/tissue identity markers are impaired from the "symmetry-breaking" periclinal cell divisions during the transition between stage I and stage II, wherein cells first acquire different identities in the proximodistal and radial axes. Particularly, PLT1, PLT2, and PLT4 genes that are typically expressed later than PLT3, PLT5, and PLT7 during LR outgrowth are not induced in the mutant primordia, rendering "PLT-null" LRP. Reintroduction of any PLT clade member in the mutant primordia completely restores layer identities at stage II and rescues mutant defects in meristem and tissue establishment. Therefore, all PLT genes can activate the formative cell divisions that lead to de novo meristem establishment and tissue patterning associated with a new growth axis.

The Roles of the Sole Activator-Type Auxin Response Factor in Pattern Formation of Marchantia polymorpha

Cell division patterning is important to determine body shape in plants. Nuclear auxin signaling mediated by AUXIN RESPONSE FACTOR (ARF) transcription factors affects plant growth and development through regulation of cell division, elongation and differentiation. The evolutionary origin of the ARF-mediated pathway dates back to at least the common ancestor of bryophytes and other land plants. The liverwort Marchantia polymorpha has three phylogenetically distinct ARFs: MpARF1, the sole 'activator' ARF; and MpARF2 and MpARF3, two 'repressor' ARFs. Genetic screens for auxin-resistant mutants revealed that loss of MpARF1 function conferred auxin insensitivity. Mparf1 mutants showed reduced auxin-inducible gene expression and various developmental defects, including thallus twisting and gemma malformation. We further investigated the role of MpARF1 in gemma development, which is traceable at the cellular level. In wild-type plants, a gemma initial first undergoes several transverse divisions to generate a single-celled stalk and a gemma proper, followed by rather synchronous longitudinal divisions in the latter. Mparf1 mutants often contained multicelled stalks and showed defects in the execution and timing of the longitudinal divisions. While wild-type gemmae finally generate two meristem notches, Mparf1 gemmae displayed various numbers of ectopic meristems. These results suggest that MpARF1 regulates formative cell divisions and axis formation through auxin responses. The mechanism for activator ARF regulation of pattern formation may be shared in land plants and therefore important for the general acquisition of three-dimensional body plans.

Brassinosteroid signaling directs formative cell divisions and protophloem differentiation in Arabidopsis root meristems.

ABSTRACTBrassinosteroids (BRs) trigger an intracellular signaling cascade through its receptors BR INSENSITIVE 1 (BRI1), BRI1-LIKE 1 (BRL1) and BRL3. Recent studies suggest that BR-independent inputs related to vascular differentiation, for instance root protophloem development, modulate downstream BR signaling components. Here, we report that protophloem sieve element differentiation is indeed impaired in bri1 brl1 brl3 mutants, although this effect might not be mediated by canonical downstream BR signaling components. We also found that their small meristem size is entirely explained by reduced cell elongation, which is, however, accompanied by supernumerary formative cell divisions in the radial dimension. Thus, reduced cell expansion in conjunction with growth retardation, because of the need to accommodate supernumerary formative divisions, can account for the overall short root phenotype of BR signaling mutants. Tissue-specific re-addition of BRI1 activity partially rescued subsets of these defects through partly cell-autonomous, partly non-cell-autonomous effects. However, protophloem-specific BRI1 expression essentially rescued all major bri1 brl1 brl3 root meristem phenotypes. Our data suggest that BR perception in the protophloem is sufficient to systemically convey BR action in the root meristem context.

Genetic and molecular mechanisms of post-embryonic root radial patterning

Arabidopsis root with its simple, consistent and well-defined pattern of cell division and growth is a highly tractable system for studying cellular development. The root apical meristem (RAM) of Arabidopsis, established post-embryonically is the main source of root cells. Complex bidirectional signaling between stem cells and mature root cells is required to maintain RAM activity. The Arabidopsis RAM comprises of a quiescent center (QC) and a niche of self-renewing stem cells that perform asymmetric (formative) cell divisions to generate daughter cells. These daughter cells subsequently divide symmetrically within delineated longitudinal cell files. The cells in the QC coordinate with neighboring stem cells to establish the balance between proliferation and differentiation in the meristem niche. The Arabidopsis primary root is composed of concentric rings of tissues (namely, the epidermis, the cortex, the endodermis and the pericycle, from the outside inwards) arranged in a radial axis that surrounds a central cylinder of vascular and procambial cells. Overlaid upon the radial organization of the root are three longitudinal zones viz, the meristem, the elongation zone and the differentiation zone, that describe cellular behavior. The boundaries of these domains are plastic and are largely regulated by antagonistic interactions between different phyto-hormones, which regulate the expression of key TFs. In this paper, we review the process of radial patterning in Arabidopsis thaliana post-embryonic root with an emphasis on genetic and molecular networks that determine the fate of each domain in the root.

Quiescent center initiation in the Arabidopsis lateral root primordia is dependent on the SCARECROW transcription factor

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.

RALFL34 regulates formative cell divisions in Arabidopsis pericycle during lateral root initiation.

In plants, many signalling molecules, such as phytohormones, miRNAs, transcription factors, and small signalling peptides, drive growth and development. However, very few small signalling peptides have been shown to be necessary for lateral root development. Here, we describe the role of the peptide RALFL34 during early events in lateral root development, and demonstrate its specific importance in orchestrating formative cell divisions in the pericycle. Our results further suggest that this small signalling peptide acts on the transcriptional cascade leading to a new lateral root upstream of GATA23, an important player in lateral root formation. In addition, we describe a role for ETHYLENE RESPONSE FACTORs (ERFs) in regulating RALFL34 expression. Taken together, we put forward RALFL34 as a new, important player in lateral root initiation.

A Feed-Forward Regulation Sets Cell Fates in Roots.

Formative cell divisions generate new cell types and tissues during development, and are controlled by receptor kinase signalling pathways. The phosphatase PP2A has now been shown to be both a target and positive regulator of the receptor kinase ACR4, thus creating a feed-forward loop that serves to establish new cell fates.

PP2A-3 interacts with ACR4 and regulates formative cell division in the Arabidopsis root

In plants, the generation of new cell types and tissues depends on coordinated and oriented formative cell divisions. The plasma membrane-localized receptor kinase ARABIDOPSIS CRINKLY 4 (ACR4) is part of a mechanism controlling formative cell divisions in the Arabidopsis root. Despite its important role in plant development, very little is known about the molecular mechanism with which ACR4 is affiliated and its network of interactions. Here, we used various complementary proteomic approaches to identify ACR4-interacting protein candidates that are likely regulators of formative cell divisions and that could pave the way to unraveling the molecular basis behind ACR4-mediated signaling. We identified PROTEIN PHOSPHATASE 2A-3 (PP2A-3), a catalytic subunit of PP2A holoenzymes, as a previously unidentified regulator of formative cell divisions and as one of the first described substrates of ACR4. Our in vitro data argue for the existence of a tight posttranslational regulation in the associated biochemical network through reciprocal regulation between ACR4 and PP2A-3 at the phosphorylation level.