Inhibition of auxin movement from the shoot into the root inhibits lateral root development in Arabidopsis

In roots two distinct polar movements of auxin have been reported that may control different developmental and growth events. To test the hypothesis that auxin derived from the shoot and transported toward the root controls lateral root development, the two polarities of auxin transport were uncoupled in Arabidopsis. Local application of the auxin-transport inhibitor naphthylphthalamic acid (NPA) at the root-shoot junction decreased the number and density of lateral roots and reduced the free indoleacetic acid (IAA) levels in the root and [3H]IAA transport into the root. Application of NPA to the basal half of or at several positions along the root only reduced lateral root density in regions that were in contact with NPA or in regions apical to the site of application. Lateral root development was restored by application of IAA apical to NPA application. Lateral root development in Arabidopsis roots was also inhibited by excision of the shoot or dark growth and this inhibition was reversible by IAA. Together, these results are consistent with auxin transport from the shoot into the root controlling lateral root development.     

Control of auxin-regulated root development by the Arabidopsis thaliana SHY2/IAA3 gene

The plant hormone auxin controls many aspects of development and acts in part by inducing expression of various genes. Arabidopsis thaliana semidominant shy2 (short hypocotyl) mutations cause leaf formation in dark-grown plants, suggesting that SHY2 has an important role in regulating development. Here we show that the SHY2 gene encodes IAA3, a previously known member of the Aux/IAA family of auxin-induced genes. Dominant shy2 mutations cause amino acid changes in domain II, conserved among all members of this family. We isolated loss-of-function shy2 alleles including a putative null mutation. Gain-of-function and loss-of-function shy2 mutations affect auxin-dependent root growth, lateral root formation, and timing of gravitropism, indicating that SHY2/IAA3 regulates multiple auxin responses in roots. The phenotypes suggest that SHY2/IAA3 may activate some auxin responses and repress others. Models invoking tissue-specificity, feedback regulation, or control of auxin transport may explain these results.     

Transport and Metabolism of the Endogenous Auxin Precursor lndole-3-Butyric Acid

T Plant growth and morphogenesis depend on the levels and distribution of the plant hormone auxin. Plants tightly regulate cellular levels of the active auxin indole-3-acetic acid (IAA) through synthesis, inactivation, and transport. Although the transporters that move IAA into and out of cells are well characterized and play important roles in development, little is known about the transport of IAA precursors. In this review, we discuss the accumulating evidence suggesting that the IAA precursor indole-3-butyric acid (IBA) is transported independently of the characterized IAA transport machinery along with the recent identification of specific IBA efflux carriers and enzymes suggested to metabolize IBA. These studies have revealed important roles for IBA in maintaining IAA levels and distribution within the plant to support normal development.     

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The plasma membrane from Arabidopsis thaliana leaves (wild-type and F mutant) has been purified by two-phase partitioning and the H(+)-transport activity was tested in vitro in the presence of different concentrations of IAA. While 1 microM of IAA had no effect, higher concentrations (10 microM and 100 microM) were inhibitory in 25 short days grown wild-type plants. However, the activity was increased by about 16% after 10 supplementary short days of growth (from 25 to 35 SD) in the represence of 10 microM of IAA. No significant sensitivity to the tested concentrations of auxin was observed during the development in short days (30, 38, 42 SD) or even the plants were induced by a 24 h of continuous light. The variability was not observed for a given developmental stage, from one experiment to another, but it was also observed after light induction of F mutant plants.     

A mutation in protein phosphatase 2A regulatory subunit A affects auxin transport in Arabidopsis

The phytohormone auxin controls processes such as cell elongation, root hair development and root branching. Tropisms, growth curvatures triggered by gravity, light and touch, are also auxin-mediated responses. Auxin is synthesized in the shoot apex and transported through the stem, but the molecular mechanism of auxin transport is not well understood. Naphthylphthalamic acid (NPA) and other inhibitors of auxin transport block tropic curvature responses and inhibit root and shoot elongation. We have isolated a novel Arabidopsis thaliana mutant designated roots curl in NPA (rcn1). Mutant seedlings exhibit altered responses to NPA in root curling and hypocotyl elongation. Auxin efflux in mutant seedlings displays increased sensitivity to NPA. The rcn1 mutation was transferred-DNA (T-DNA) tagged and sequences flanking the T-DNA insert were cloned. Analysis of the RCN1 cDNA reveals that the T-DNA insertion disrupts a gene for the regulatory A subunit of protein phosphatase 2A (PP2A-A). The RCN1 gene rescues the rcn1 mutant phenotype and also complements the temperature-sensitive phenotype of the Saccharomyces cerevisiae PP2A-A mutation, tpd3-1. These data implicate protein phosphatase 2A in the regulation of auxin transport in Arabidopsis.     

EIR1, a root-specific protein involved in auxin transport, is required for gravitropism in Arabidopsis thaliana

The EIR1 gene of Arabidopsis is a member of a family of plant genes with similarities to bacterial membrane transporters. This gene is expressed only in the root, which is consistent with the phenotypes of the eir1 mutants-the roots are agravitropic and have a reduced sensitivity to ethylene. The roots of eir1 mutants are also insensitive to the excess auxin produced by alf1-1 and fail to induce an auxin-inducible gene in the expansion zone. Although they fail to respond to internally generated auxin, they respond normally to externally applied auxin. Expression of the EIR1 gene in Saccharomyces cerevisiae confers resistance to fluorinated indolic compounds. Taken together, these data suggest that the EIR1 protein has a root-specific role in the transport of auxin.     

Arabidopsis AUX1 gene: a permease-like regulator of root gravitropism

The plant hormone auxin regulates various developmental processes including root formation, vascular development, and gravitropism. Mutations within the AUX1 gene confer an auxin-resistant root growth phenotype and abolish root gravitropic curvature. Polypeptide sequence similarity to amino acid permeases suggests that AUX1 mediates the transport of an amino acid-like signaling molecule. Indole-3-acetic acid, the major form of auxin in higher plants, is structurally similar to tryptophan and is a likely substrate for the AUX1 gene product. The cloned AUX1 gene can restore the auxin-responsiveness of transgenic aux1 roots. Spatially, AUX1 is expressed in root apical tissues that regulate root gravitropic curvature.     

Plant hormones: ins and outs of auxin transport

Regulated transport has long been known to play a key part in action of the plant hormone auxin. Now, at last, a family of auxin efflux carriers has been identified, and the characterisation of one family member has provided strong evidence in support of models that have been proposed to explain gravitropic curvature in roots.     

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Mutations in the Agr locus of Arabidopsis thaliana impair the root gravitropic response. Root growth of agr mutants is moderately resistant to ethylene and to an auxin transport inhibitor. Vertically placed agr roots grow into agar medium containing IAA or naphthalene-1-acetic acid, but not into medium containing 2,4-D. Positional cloning showed that AGR encodes a root-specific member of a novel membrane-protein family with limited homology to bacterial transporters.     

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Polar auxin transport plays a key role in the regulation of plant growth and development. To identify genes involved in this process, we have developed a genetic procedure to screen for mutants of Arabidopsis that are altered in their response to auxin transport inhibitors. We recovered a total of 16 independent mutants that defined seven genes, called TRANSPORT INHIBITOR RESPONSE (TIR) genes. Recessive mutations in one of these genes, TIR3, result in altered responses to transport inhibitors, a reduction in polar auxin transport, and a variety of morphological defects that can be ascribed to changes in indole-3-acetic acid distribution. Most dramatically, tir3 seedlings are strongly deficient in lateral root production, a process that is known to depend on polar auxin transport from the shoot into the root. In addition, tir3 plants display a reduction in apical dominance as well as decreased elongation of siliques, pedicels, roots, and the inflorescence. Biochemical studies indicate that tir3 plants have a reduced number of N-1-naphthylphthalamic (NPA) binding sites, suggesting that the TIR3 gene is required for expression, localization, or stabilization of the NPA binding protein (NBP). Alternatively, the TIR3 gene may encode the NBP. Because the tir3 mutants have a substantial defect in NPA binding, their phenotype provides genetic evidence for a role for the NBP in plant growth and development.     

Molecular cloning and expression of the early auxin-responsive Aux/IAA gene family in Nicotiana tabacum

Early auxin-regulated tobacco cDNAs, belonging to the Aux/IAA gene family have been isolated by screening of a cDNA library prepared from auxin-treated suspension-grown etiolated seedlings of Nicotiana tabacum. The probes used were either RT-PCR fragments or an insert resulting from mRNA differential display selection. All of them possessed the structural features which characterize the Aux/IAA gene products. The auxin response of three distinct Nt-iaa subclasses has been characterized in terms of kinetics, dose-response and specificity as several plant hormones and chemicals have been tested for their ability to alter Nt-iaa mRNA accumulation. Differences of auxin responses have been observed between the Nt-iaa analysed, revealing significant differences of regulation. The effect of the protein synthesis inhibitor cycloheximide suggested that Nt-iaa2.3, Nt-iaa4.3 and strictly related genes can be classified as primary auxin-responsive genes and Nt-iaa28 as a late one. The steady-state mRNA level of these Nt-iaa has also been determined in organs of tobacco plants.     

Arabidopsis TCH4, regulated by hormones and the environment, encodes a xyloglucan endotransglycosylase

Adaptation of plants to environmental conditions requires that sensing of external stimuli be linked to mechanisms of morphogenesis. The Arabidopsis TCH (for touch) genes are rapidly upregulated in expression in response to environmental stimuli, but a connection between this molecular response and developmental alterations has not been established. We identified TCH4 as a xyloglucan endotransglycosylase by sequence similarity and enzyme activity. Xyloglucan endotransglycosylases most likely modify cell walls, a fundamental determinant of plant form. We determined that TCH4 expression is regulated by auxin and brassinosteroids, by environmental stimuli, and during development, by a 1-kb region. Expression was restricted to expanding tissues and organs that undergo cell wall modification. Regulation of genes encoding cell wall-modifying enzymes, such as TCH4, may underlie plant morphogenetic responses to the environment.     

The arabidopsis ACT7 actin gene is expressed in rapidly developing tissues and responds to several external stimuli

ACT7 encodes one of the six distinct and ancient subclasses of actin protein in the complex Arabidopsis actin gene family. We determined the sequence and structure of the Arabidopsis thaliana ACT7 actin gene and investigated its tissue-specific expression and regulation. The ACT7 mRNA levels varied by 128-fold among several different tissues and organs. The highest levels of aCT7 mRNA were found in rapidly expanding vegetative organs, the lowest in pollen. A translational fusion with the 5' end of ACT 7 (1.9 kb) joined to the beta-glucuronidase reporter gene was strongly and preferentially expressed in all young, developing vegetative tissues of transgenic Arabidopsis plants. ACT7 was the only Arabidopsis actin gene strongly expressed in the hypocotyl and seed coat. Although no beta-glucuronidase expression was seen in developing ovules or immature seeds, strong expression was seen in dry seeds and immediately after imbibition in the entire seedling. ACT7 was the only Arabidopsis actin gene to respond strongly to auxin, other hormone treatments, light regime, and wounding, and may be the primary actin gene responding to external stimuli. The ACT7 promoter sequence contains a remarkable number of motifs with sequence similarity to putative phytohormone response elements.     

Cavoportal collateral pathways in vena cava obstruction: imaging features

The vascular tissues of flowering plants form networks of interconnected cells throughout the plant body. The molecular mechanisms directing the routes of vascular strands and ensuring tissue continuity within the vascular system are not known, but are likely to depend on general cues directing plant cell orientation along the apical-basal axis. Mutations in the Arabidopsis gene MONOPTEROS (MP) interfere with the formation of vascular strands at all stages and also with the initiation of the body axis in the early embryo. Here we report the isolation of the MP gene by positional cloning. The predicted protein product contains functional nuclear localization sequences and a DNA binding domain highly similar to a domain shown to bind to control elements of auxin inducible promoters. During embryogenesis, as well as organ development, MP is initially expressed in broad domains that become gradually confined towards the vascular tissues. These observations suggest that the MP gene has an early function in the establishment of vascular and body patterns in embryonic and post-embryonic development.     

A major isoform of the maize plasma membrane H(+)-ATPase: characterization and induction by auxin in coleoptiles

The plasma membrane (PM) H(+)-ATPase has been proposed to play important transport and regulatory roles in plant physiology, including its participation in auxin-induced acidification in coleoptile segments. This enzyme is encoded by a family of genes differing in tissue distribution, regulation, and expression level. A major expressed isoform of the maize PM H(+)-ATPase (MHA2) has been characterized. RNA gel blot analysis indicated that MHA2 is expressed in all maize organs, with highest levels being in the roots. In situ hybridization of sections from maize seedlings indicated enriched expression of MHA2 in stomatal guard cells, phloem cells, and root epidermal cells. MHA2 mRNA was induced threefold when nonvascular parts of the coleoptile segments were treated with auxin. This induction correlates with auxin-triggered proton extrusion by the same part of the segments. The PM H(+)-ATPase in the vascular bundies does not contribute significantly to auxin-induced acidification, is not regulated by auxin, and masks the auxin effect in extracts of whole coleoptile segments. We conclude that auxin-induced acidification in coleoptile segments most often occurs in the nonvascular tissue and is mediated, at least in part, by increased levels of MHA2.     

Seven Things We Think We Know about Auxin Transport

T Polar transport of the phytohormone auxin and the establishment of localized auxin maxima regulate embryonic development, stem cell maintenance, root and shoot architecture, and tropic growth responses. The past decade has been marked by dramatic progress in efforts to elucidate the complex mechanisms by which auxin transport regulates plant growth. As the understanding of auxin transport regulation has been increasingly elaborated, it has become clear that this process is involved in almost all plant growth and environmental responses in some way. However, we still lack information about some basic aspects of this fundamental regulatory mechanism. In this review, we present what we know (or what we think we know) and what we do not know about seven auxin-regulated processes. We discuss the role of auxin transport in gravitropism in primary and lateral roots, phototropism, shoot branching, leaf expansion, and venation. We also discuss the auxin reflux/fountain model at the root tip, flavonoid modulation of auxin transport processes, and outstanding aspects of post-translational regulation of auxin transporters. This discussion is not meant to be exhaustive, but highlights areas in which generally held assumptions require more substantive validation.