The auxin-insensitive bodenlos mutation affects primary root formation and apical-basal patterning in the Arabidopsis embryo.
In Arabidopsis embryogenesis, the primary root meristem originates from descendants of both the apical and the basal daughter cell of the zygote. We have isolated a mutant of a new gene named BODENLOS (BDL) in which the primary root meristem is not formed whereas post-embryonic roots develop and bdl seedlings give rise to fertile adult plants. Some bdl seedlings lacked not only the root but also the hypocotyl, thus resembling monopteros (mp) seedlings. In addition, bdl seedlings were insensitive to the auxin analogue 2,4-D, as determined by comparison with auxin resistant1 (axr1) seedlings. bdl embryos deviated from normal development as early as the two-cell stage at which the apical daughter cell of the zygote had divided horizontally instead of vertically. Subsequently, the uppermost derivative of the basal daughter cell, which is normally destined to become the hypophysis, divided abnormally and failed to generate the quiescent centre of the root meristem and the central root cap. We also analysed double mutants. bdl mp embryos closely resembled the two single mutants, bdl and mp, at early stages, while bdl mp seedlings essentially consisted of hypocotyl but did form primary leaves. bdl axr1 embryos approached the mp phenotype at later stages, and bdl axr1 seedlings resembled mp seedlings. Our results suggest that BDL is involved in auxin-mediated processes of apical-basal patterning in the Arabidopsis embryo. (+info)
The DNA binding site of the Dof protein NtBBF1 is essential for tissue-specific and auxin-regulated expression of the rolB oncogene in plants.
The Dof proteins are a large family of plant transcription factors that share a single highly conserved zinc finger. The tobacco Dof protein NtBBF1 was identified by its ability to bind to regulatory domain B in the promoter of the rolB oncogene. In this study, we show that the ACT T TA target sequence of NtBBF1 in domain B is necessary for tissue-specific expression of rolB. beta-Glucuronidase (GUS) activity of tobacco plants containing a rolB promoter-GUS fusion with a mutated NtBBF1 target sequence within domain B is almost completely suppressed in apical meristems and is severely abated in the vascular system. The ACT T TA motif is shown here also to be one of the cis-regulatory elements involved in auxin induction of rolB. The pattern of NtBBF1 expression in plants is remarkably similar to that of rolB, except in mesophyll cells of mature leaves, in which only NtBBF1 expression could be detected. Ectopic expression of rolB in mesophyll cells was achieved by particle gun delivery if the NtBBF1 binding sequence was intact. These data provide evidence that in the plant, a Dof protein DNA binding sequence acts as a transcriptional regulatory motif, and they point to NtBBF1 as the protein involved in mediating tissue-specific and auxin-inducible expression of rolB. (+info)
IAR3 encodes an auxin conjugate hydrolase from Arabidopsis.
Amide-linked conjugates of indole-3-acetic acid (IAA) are putative storage or inactivation forms of the growth hormone auxin. Here, we describe the Arabidopsis iar3 mutant that displays reduced sensitivity to IAA-Ala. IAR3 is a member of a family of Arabidopsis genes related to the previously isolated ILR1 gene, which encodes an IAA-amino acid hydrolase selective for IAA-Leu and IAA-Phe. IAR3 and the very similar ILL5 gene are closely linked on chromosome 1 and comprise a subfamily of the six Arabidopsis IAA-conjugate hydrolases. The purified IAR3 enzyme hydrolyzes IAA-Ala in vitro. iar 3 ilr1 double mutants are more resistant than either single mutant to IAA-amino acid conjugates, and plants overexpressing IAR3 or ILR1 are more sensitive than is the wild type to certain IAA-amino acid conjugates, reflecting the overlapping substrate specificities of the corresponding enzymes. The IAR3 gene is expressed most strongly in roots, stems, and flowers, suggesting roles for IAA-conjugate hydrolysis in those tissues. (+info)
Biosynthesis of indole-3-acetic acid in Azospirillum brasilense. Insights from quantum chemistry.
Quantum chemical methods AM1 and PM3 and chromatographic methods were used to qualitatively characterize pathways of bacterial production of indole-3-acetic acid (IAA). The standard free energy changes (delta G(o)'sum) for the synthesis of tryptophan (Trp) from chorismic acid via anthranilic acid and indole were calculated, as were those for several possible pathways for the synthesis of IAA from Trp, namely via indole-3-acetamide (IAM), indole-3-pyruvic acid (IPyA), and indole-3-acetonitrile (IAN). The delta G(o)'sum for Trp synthesis from chorismic acid was -402 (-434) kJ.mol-1 (values in parentheses were calculated by PM3). The delta G(o)'sum for IAA synthesis from Trp were -565 (-548) kJ.mol-1 for the IAN pathway, -481 (-506) kJ.mol-1 for the IAM pathway, and -289 (-306) kJ.mol-1 for the IPyA pathway. By HPLC analysis, the possibility was assessed that indole, anthranilic acid, and Trp might be utilized as precursors for IAA synthesis by Azospirillum brasilense strain Sp 245. The results indicate that there is a high motive force for Trp synthesis from chorismic acid and for IAA synthesis from Trp, and make it unlikely that anthranilic acid and indole act as the precursors to IAA in a Trp-independent pathway. (+info)
Reconstitution of an electrogenic auxin transport activity mediated by Arabidopsis thaliana plasma membrane proteins.
Plasma membrane proteins from Arabidopsis thaliana leaves were reconstituted into proteoliposomes and a K+ diffusion potential was generated. The resulting ionic fluxes, determined in the presence of the plant hormone auxin (indole-3 acetic acid), showed an additional electrogenic and saturable component, with a K(M) of 6 microM. This flux was neither detected in liposomes in the presence of indole-3 acetic acid, nor in proteoliposomes in the presence of an inactive auxin analog and was completely inhibited by 3 microM naphtylphthalamic acid, a specific inhibitor of the auxin efflux carrier. The efficiency of the reconstituted carrier and the mechanism of its regulation by naphtylphthalamic acid are discussed. (+info)
Novel gating mechanism of polyamine block in the strong inward rectifier K channel Kir2.1.
Inward rectifying K channels are essential for maintaining resting membrane potential and regulating excitability in many cell types. Previous studies have attributed the rectification properties of strong inward rectifiers such as Kir2.1 to voltage-dependent binding of intracellular polyamines or Mg to the pore (direct open channel block), thereby preventing outward passage of K ions. We have studied interactions between polyamines and the polyamine toxins philanthotoxin and argiotoxin on inward rectification in Kir2.1. We present evidence that high affinity polyamine block is not consistent with direct open channel block, but instead involves polyamines binding to another region of the channel (intrinsic gate) to form a blocking complex that occludes the pore. This interaction defines a novel mechanism of ion channel closure. (+info)
Glutamate receptor expression regulates quantal size and quantal content at the Drosophila neuromuscular junction.
At the Drosophila glutamatergic neuromuscular junction, the postsynaptic cell can regulate synaptic strength by both changing its sensitivity to neurotransmitter and generating a retrograde signal that regulates presynaptic transmitter release. To investigate the molecular mechanisms underlying these forms of plasticity, we have undertaken a genetic analysis of two postsynaptic glutamate receptors that are expressed at this synapse. Deletion of both genes results in embryonic lethality that can be rescued by transgenic expression of either receptor. Although these receptors are redundant for viability, they have important differences. By transgenically rescuing the double mutant, we have investigated the relationship of receptor gene dosage and composition to synaptic function. We find that the receptor subunit composition regulates quantal size, Argiotoxin sensitivity, and receptor desensitization kinetics. Finally, we show that the activity of the receptor can regulate the retrograde signal functioning at this synapse. Thus, the diversity of receptors expressed at this synapse provides the cell with mechanisms for generating synaptic plasticity. (+info)
AUX1 regulates root gravitropism in Arabidopsis by facilitating auxin uptake within root apical tissues.
Plants employ a specialized transport system composed of separate influx and efflux carriers to mobilize the plant hormone auxin between its site(s) of synthesis and action. Mutations within the permease-like AUX1 protein significantly reduce the rate of carrier-mediated auxin uptake within Arabidopsis roots, conferring an agravitropic phenotype. We are able to bypass the defect within auxin uptake and restore the gravitropic root phenotype of aux1 by growing mutant seedlings in the presence of the membrane-permeable synthetic auxin, 1-naphthaleneacetic acid. We illustrate that AUX1 expression overlaps that previously described for the auxin efflux carrier, AtPIN2, using transgenic lines expressing an AUX1 promoter::uidA (GUS) gene. Finally, we demonstrate that AUX1 regulates gravitropic curvature by acting in unison with the auxin efflux carrier to co-ordinate the localized redistribution of auxin within the Arabidopsis root apex. Our results provide the first example of a developmental role for the auxin influx carrier within higher plants and supply new insight into the molecular basis of gravitropic signalling. (+info)