A role for gibberellic acid in orienting microtubules and regulating cell growth polarity in the maize root cortex

The role of gibberellins and cortical microtubules in determining the polarity of cell growth in the root cortex of maize (Zea mays L.) was examined. Inhibition of gibberellin biosynthesis, either naturally through mutation (d5 mutant) or by means of chemicals such as 2S,3S paclobutrazol, caused thickening of root apices and increased their starch content. Immunofluorescence microscopy of cortical microtubules, coupled with a comparison of cell widhts, lengths and shapes, indicated that the meristem and immediate post-mitotic zone were the targets of gibberellin deficiency. Cortical cells in these regions were impaired in their ability to develop highly ordered transversal arrays of cortical microtubules. Consequently, the cells became wider and shorter. Application of gibberellic acid re-established the arrangements of cortical microtubules and the polarity of cell growth characteristic for roots having normal levels of gibberellins, it also decreased the starch content. These results indicate that gibberellins are morphogenetically active substances, not only in shoots but also in roots of maize.Abbreviations CMT cortical microtubule - GA gibberellin - GA₃ gibberellic acid - MT microtubule - PIG postmitotic isodiametric growth The authors acknowledge the support to F.B. from the Royal Society (London UK). We also thank Dr. J. Lenton (University of Bristol, Long Ashton Research Station) who kindly supplied us with 2S,3S paclobutrazol and grains of the GA-deficient d5 mutant of maize.     

VAP-RELATED SUPPRESSORS OF TOO MANY MOUTHS (VST) family proteins are regulators of root system architecture

Root system architecture (RSA) is a key factor in the efficiency of nutrient capture and water uptake in plants. Understanding the genetic control of RSA will be useful in minimizing fertilizer and water usage in agricultural cropping systems. Using a hydroponic screen and a gel-based imaging system, we identified a rice (Oryza sativa) gene, VAP-RELATED SUPPRESSOR OF TOO MANY MOUTHS1 (OsVST1), which plays a key role in controlling RSA. This gene encodes a homolog of the VAP-RELATED SUPPRESSORS OF TOO MANY MOUTHS (VST) proteins in Arabidopsis (Arabidopsis thaliana), which promote signaling in stomata by mediating plasma membrane–endoplasmic reticulum contacts. OsVST1 mutants have shorter primary roots, decreased root meristem size, and a more compact RSA. We show that the Arabidopsis VST triple mutants have similar phenotypes, with reduced primary root growth and smaller root meristems. Expression of OsVST1 largely complements the short root length and reduced plant height in the Arabidopsis triple mutant, supporting conservation of function between rice and Arabidopsis VST proteins. In a field trial, mutations in OsVST1 did not adversely affect grain yield, suggesting that modulation of this gene could be used as a way to optimize RSA without an inherent yield penalty.

Root meristem size and root system architecture in both rice and Arabidopsis are regulated by proteins related to mediators plasma membrane–endoplasmic reticulum contact.     

Arabidopsis CULLIN3 Genes Regulate Primary Root Growth and Patterning by Ethylene-Dependent and -Independent Mechanisms

CULLIN3 (CUL3) together with BTB-domain proteins form a class of Cullin-RING ubiquitin ligases (called CRL3s) that control the rapid and selective degradation of important regulatory proteins in all eukaryotes. Here, we report that in the model plant Arabidopsis thaliana, CUL3 regulates plant growth and development, not only during embryogenesis but also at post-embryonic stages. First, we show that CUL3 modulates the emission of ethylene, a gaseous plant hormone that is an important growth regulator. A CUL3 hypomorphic mutant accumulates ACS5, the rate-limiting enzyme in ethylene biosynthesis and as a consequence exhibits a constitutive ethylene response. Second, we provide evidence that CUL3 regulates primary root growth by a novel ethylene-dependant pathway. In particular, we show that CUL3 knockdown inhibits primary root growth by reducing root meristem size and cell number. This phenotype is suppressed by ethylene-insensitive or resistant mutations. Finally, we identify a function of CUL3 in distal root patterning, by a mechanism that is independent of ethylene. Thus, our work highlights that CUL3 is essential for the normal division and organisation of the root stem cell niche and columella root cap cells.     

Arabidopsis ASA1 Is Important for Jasmonate-Mediated Regulation of Auxin Biosynthesis and Transport during Lateral Root Formation

Plant roots show an impressive degree of plasticity in adapting their branching patterns to ever-changing growth conditions. An important mechanism underlying this adaptation ability is the interaction between hormonal and developmental signals. Here, we analyze the interaction of jasmonate with auxin to regulate lateral root (LR) formation through characterization of an Arabidopsis thaliana mutant, jasmonate-induced defective lateral root1 (jdl1/asa1-1). We demonstrate that, whereas exogenous jasmonate promotes LR formation in wild-type plants, it represses LR formation in jdl1/asa1-1. JDL1 encodes the auxin biosynthetic gene ANTHRANILATE SYNTHASE α1 (ASA1), which is required for jasmonate-induced auxin biosynthesis. Jasmonate elevates local auxin accumulation in the basal meristem of wild-type roots but reduces local auxin accumulation in the basal meristem of mutant roots, suggesting that, in addition to activating ASA1-dependent auxin biosynthesis, jasmonate also affects auxin transport. Indeed, jasmonate modifies the expression of auxin transport genes in an ASA1-dependent manner. We further provide evidence showing that the action mechanism of jasmonate to regulate LR formation through ASA1 differs from that of ethylene. Our results highlight the importance of ASA1 in jasmonate-induced auxin biosynthesis and reveal a role for jasmonate in the attenuation of auxin transport in the root and the fine-tuning of local auxin distribution in the root basal meristem.     

Activation of gibberellin 20-oxidase 2 undermines auxin-dependent root and root hair growth in NaCl-stressed <Emphasis Type="Italic">Arabidopsis</Emphasis> seedlings

Although salt stress mainly disturbs plant root growth by affecting the biosynthesis and signaling of phytohormones, such as gibberellin (GA) and auxin, the exact mechanisms of the crosstalk between these two hormones remain to be clarified. Indole-3-acetic acid (IAA) is a biologically active auxin molecule. In this study, we investigated the role of Arabidopsis GA20-oxidase 2 (GA20ox2), a final rate-limiting enzyme of active GA biosynthesis, in IAA-directed root growth under NaCl stress. Under the NaCl treatment, seedlings of a loss-of-function ga20ox2-1 mutant exhibited primary root and root hair elongation, altered GA₄ accumulation, and decreased root Na⁺ contents compared with the wild-type, transgenic GA20ox2-complementing, and GA20ox2-overexpression plant lines. Concurrently, ga20ox2-1 alleviated the tissue-specific inhibition of NaCl on IAA generation by YUCCAs, IAA transport by PIN1 and PIN2, and IAA accumulation in roots, thereby explaining how NaCl increased GA20ox2 expression in shoots but disrupted primary root and root hair growth in wild-type seedlings. In addition, a loss-of-function pin2 mutant impeded GA20ox2 expression, indicating that GA20ox2 function requires PIN2 activity. Thus, the activation of GA20ox2 retards IAA-directed primary root and root hair growth in response to NaCl stress.     

DAR2 acts as an important node connecting cytokinin,auxin, SHY2 and PLT1/2 in root meristem size control

Cytokinin and auxin antagonistically affect cell proliferation and differentiation and thus regulate root meristem size by influencing the abundance of SHORT HYPOCOTYL2 (SHY2/IAA3). SHY2 affects auxin distribution in the root meristem by repressing the auxin-inducible expression of PIN-FORMED (PIN) auxin transport genes. The PLETHORA (PLT1/2) genes influence root meristem growth by promoting stem cells and transit-amplifying cells. However, the factors connecting cytokinin, auxin, SHY2 and PLT1/2 are largely unknown. In a recent study, we have shown that the DA1-related protein 2 (DAR2) acts downstream of cytokinin and SHY2 but upstream of PLT1/2 to affect root meristem size. Here, we discuss the possible molecular mechanisms by which Arabidopsis DAR2 controls root meristem size.     

Gibberellin disturbs the balance of endogenesis hormones and inhibits adventitious root development of Pseudostellaria heterophylla through regulating gene expression related to hormone synthesis

The adventitious roots of some plants will develop into tuberous roots which are widely used in many traditional Chinese medicines, including Pseudostellaria heterophylla. If adventitious root development is inhibited, the yield of Chinese medicinal materials will be reduced. Gibberellic acid is an important phytohormone that promotes plant growth and increases the resistance to drought, flood or disease. However, the effects of gibberellic acid on adventitious roots of Pseudostellaria heterophylla are not clear. Here, we reports GA3 suppressed adventitious root development of Pseudostellaria heterophylla by disturbing the balance of endogenesis hormones. By detecting the contents of various endogenous hormones, we found that the development of adventitious roots negatively correlated with the content of CA3 in tuberous roots. Exogenous GA3 treatment decreased the diameter of adventitious roots, but increased the length of adventitious roots of Pseudostellaria heterophylla. In contrast, blocking the biosynthesis of GA3 suppressed stem growth and promoted the xylem of tuberous roots development. Moreover, exogenous GA3 treatment resulted in imbalance of endogenesis hormones by regulating their synthesis-related genes expression in xylem of tuberous roots. These results suggest GA3 broke the established distribution of hormones by regulating synthesis, transport and biological activation of hormones to activate the apical meristem and suppress lateral meristem. Regulating GA3 signaling during adventitious roots development would be one of the possible ways to increase the yield of P. heterophylla.     

Internodal elongation and orientation of cellulose microfibrils and microtubules in deepwater rice

Excised stem sections of deepwater rice (Oryza sativa L.) containing the highest internode were used to study the induction of rapid internodal elongation by gibberellin (GA). It has been shown before that this growth response is based on enhanced cell division in the intercalary meristem and on increased cell elongation. In both GA-treated and control stem sections, the basal 5-mm region of the highest internode grows at the fastest rate. During 24 h of GA treatment, the internodal elongation zone expands from 15 to 35 mm. Gibberellin does not promote elongation of internodes from which the intercalary meristem has been excised. The orientation of cellulose microfibrils (CMFs) is a determining factor in cell growth. Elongation is favored when CMFs are oriented transversely to the direction of growth while elongation is limited when CMFs are oriented in the oblique or longitudinal direction. The orientation of CMFs in parenchymal cells of GA-treated and control internodes is transverse throughout the internode, indicating that CMFs do not restrict elongation of these cells. Changes in CMF orientation were observed in epidermal cells, however. In the basal 5-mm zone of the internode, which includes the intercalary meristem, CMFs of the epidermal cell walls are transversely oriented in both GA-treated and control stem sections. In slowly growing control internodes, CMF orientation changes to the oblique as cells are displaced from this basal 5-mm zone to the region above it. In GA-treated rapidly growing internodes, the reorientation of CMFs from the transverse to the oblique is more gradual and extends over the 35-mm length of the elongation zone. The CMFs of older epidermal cells are obliquely oriented in control and GA-treated internodes. The orientation of the CMFs parallels that of the cortical microtubules. This is consistent with the hypothesis that cortical microtubules determine the direction of CMF deposition. We conclude that GA acts on cells that have transversely oriented CMFs but does not promote growth of cells whose CMFs are already obliquely oriented at the start of GA treatment.     

The ROOT MERISTEMLESS1/CADMIUM SENSITIVE2 gene defines a glutathione-dependent pathway involved in initiation and maintenance of cell division during postembryonic root development

Activation of cell division in the root apical meristem after germination is essential for postembryonic root development. Arabidopsis plants homozygous for a mutation in the ROOT MERISTEMLESS1 (RML1) gene are unable to establish an active postembryonic meristem in the root apex. This mutation abolishes cell division in the root but not in the shoot. We report the molecular cloning of the RML1 gene, which encodes the first enzyme of glutathione (GSH) biosynthesis, γ-glutamylcysteine synthetase, and which is allelic to CADMIUM SENSITIVE2. The phenotype of the rml1 mutant, which was also evident in the roots of wild-type Arabidopsis and tobacco treated with an inhibitor of GSH biosynthesis, could be relieved by applying GSH to rml1 seedlings. By using a synchronized tobacco cell suspension culture, we showed that the G₁-to-S phase transition requires an adequate level of GSH. These observations suggest the existence of a GSH-dependent developmental pathway essential for initiation and maintenance of cell division during postembryonic root development.     

Grafting analysis indicates that malfunction of TRICOT in the root causes a nodulation-deficient phenotype in Lotus japonicus

Leguminous plants develop root nodules in symbiosis with soil rhizobia. Nodule formation occurs following rhizobial infection of the host root that induces dedifferentiation of some cortical cells and the initiation of a new developmental program to form nodule primordia. In a recent study, we identified a novel gene, TRICOT (TCO), that acts as a positive regulator of nodulation in Lotus japonicus. In addition to its role in nodulation, tco mutant plants display pleiotropic defects including abnormal shoot apical meristem formation. Here, we investigated the effect of the tco mutation on nodulation using a grafting approach. The results strongly indicate that the nodulation-deficient phenotype of the mutant results from malfunction of the TCO gene in the root.     

Proteome approach to characterize the methylmalonate-semialdehyde dehydrogenase that is regulated by gibberellin

Proteins regulated by gibberellin (GA) in rice were determined by proteome analysis. Proteins extracted from suspension culture cells of slr1, a constitutive GA response mutant of rice, were separated by two-dimensional polyacrylamide gel electrophoresis, and three proteins were greatly accumulated in the mutant. The most up-regulated protein was methylmalonate-semialdehyde dehydrogenase (MMSDH), and the amount of protein was 7-fold that of wild type. In this study, the function of MMSDH in rice was analyzed. MMSDH gene expression in suspension culture cells, roots, and leaf sheaths ofslr1 was higher than that in its wild-type. MMSDH expression in wild-type roots was increased by exogenous GA(3). Analyzed by in situ hybridization, MMSDH mRNA was expressed in root primordia of slr1, where cells are undergoing growth. MMSDH gene expression in the root zone of tissue differentiation was higher than in the elongation zone or meristem. Transgenic rice expressing antisense MMSDH showed that its seminal roots were thinner than that of control, and that the leaf sheath elongation was slightly inhibited compared to control. Concentrations of TCA cycle metabolites were decreased in the antisense plants as compared with the control plants, suggesting that acetyl-CoA was reduced in the antisense plants. These results suggest that one of the regulations by GA signal transduction including SLR1 is the expression of MMSDH, and that MMSDH may play a role in root development and leaf sheath elongation in rice.     

Tall or short? Slender or thick? A plant strategy for regulating elongation growth of roots by low concentrations of gibberellin

Background

Since the plant hormone gibberellin (GA) was discovered as a fungal toxin that caused abnormal elongation of rice shoots, the physiological function of GA has mainly been investigated in relation to the regulation of plant height. However, an indispensable role for GA in root growth has been elucidated by using severely GA-depleted plants, either with a gene mutation in GA biosynthesis or which have been treated by an inhibitor of GA biosynthesis. The molecular sequence of GA signalling has also been studied to understand GA functions in root growth.

Scope

This review addresses research progress on the physiological functions of GA in root growth. Concentration-dependent stimulation of elongation growth by GA is important for the regulation of plant height and root length. Thus the endogenous level of GA and/or the GA sensitivity of shoots and roots plays a role in determining the shoot-to-root ratio of the plant body. Since the shoot-to-root ratio is an important parameter for agricultural production, control of GA production and GA sensitivity may provide a strategy for improving agricultural productivity. The sequence of GA signal transduction has recently been unveiled, and some component molecules are suggested as candidate in planta regulatory sites and as points for the artificial manipulation of GA-mediated growth control.

Conclusions

This paper reviews: (1) the breakthrough dose–response experiments that show that root growth is regulated by GA in a lower concentration range than is required for shoot growth; (2) research on the regulation of GA biosynthesis pathways that are known predominantly to control shoot growth; and (3) recent research on GA signalling pathways, including GA receptors, which have been suggested to participate in GA-mediated growth regulation. This provides useful information to suggest a possible strategy for the selective control of shoot and root growth, and to explain how GA plays a role in rosette and liana plants with tall or short, and slender or thick axial organs.     

Phenotype of Arabidopsis thaliana semi-dwarfs with deep roots and high growth rates under water-limiting conditions is independent of the GA5 loss-of-function alleles

Background and Aims The occurrence of Arabidopsis thaliana semi-dwarf accessions carrying inactive alleles at the gibberellin (GA) biosynthesis GA5 locus has raised the question whether there are pleiotropic effects on other traits at the root level, such as rooting depth. In addition, it is unknown whether semi-dwarfism in arabidopsis confers a growth advantage under water-limiting conditions compared with wild-type plants. The aim of this research was therefore to investigate whether semi-dwarfism has a pleiotropic effect in the root system and also whether semi-dwarfs might be more tolerant of water-limiting conditions.Methods The root systems of different arabidopsis semi-dwarfs and GA biosynthesis mutants were phenotyped in vitro using the GROWSCREEN-ROOT image-based software. Semi-dwarfs were phenotyped together with tall, near-related accessions. In addition, root phenotypes were investigated in soil-filled rhizotrons. Rosette growth trajectories were analysed with the GROWSCREEN-FLUORO setup based on non-invasive imaging.Key Results Mutations in the early steps of the GA biosynthesis pathway led to a reduction in shoot as well as root size. Depending on the genetic background, mutations at the GA5 locus yielded phenotypes characterized by decreased root length in comparison with related wild-type ones. The semi-dwarf accession Pak-3 showed the deepest root system both in vitro and in soil cultivation experiments; this comparatively deep root system, however, was independent of the ga5 loss-of-function allele, as shown by co-segregation analysis. When the accessions were grown under water-limiting conditions, semi-dwarf accessions with high growth rates were identified.Conclusions The observed diversity in root system growth and architecture occurs independently of semi-dwarf phenotypes, and is probably linked to a genetic background effect. The results show that there are no clear advantages of semi-dwarfism at low water availability in arabidopsis.