An auxin-dependent distal organizer of pattern and polarity in the Arabidopsis root.
(33/2030)
Root formation in plants involves the continuous interpretation of positional cues. Physiological studies have linked root formation to auxins. An auxin response element displays a maximum in the Arabidopsis root and we investigate its developmental significance. Auxin response mutants reduce the maximum or its perception, and interfere with distal root patterning. Polar auxin transport mutants affect its localization and distal pattern. Polar auxin transport inhibitors cause dramatic relocalization of the maximum, and associated changes in pattern and polarity. Auxin application and laser ablations correlate root pattern with a maximum adjacent to the vascular bundle. Our data indicate that an auxin maximum at a vascular boundary establishes a distal organizer in the root. (+info)
Auxin is required for leaf vein pattern in Arabidopsis.
(34/2030)
To investigate possible roles of polar auxin transport in vein patterning, cotyledon and leaf vein patterns were compared for plants grown in medium containing polar auxin transport inhibitors (N-1-naphthylphthalamic acid, 9-hydroxyfluorene-9-carboxylic acid, and 2,3,5-triiodobenzoic acid) and in medium containing a less well-characterized inhibitor of auxin-mediated processes, 2-(p-chlorophynoxy)-2-methylpropionic acid. Cotyledon vein pattern was not affected by any inhibitor treatments, although vein morphology was altered. In contrast, leaf vein pattern was affected by inhibitor treatments. Growth in polar auxin transport inhibitors resulted in leaves that lacked vascular continuity through the petiole and had broad, loosely organized midveins, an increased number of secondary veins, and a dense band of misshapen tracheary elements adjacent to the leaf margin. Analysis of leaf vein pattern developmental time courses suggested that the primary vein did not develop in polar auxin transport inhibitor-grown plants, and that the broad midvein observed in these seedlings resulted from the coalescence of proximal regions of secondary veins. Possible models for leaf vein patterning that could account for these observations are discussed. (+info)
An expansin gene expressed in ripening strawberry fruit.
(35/2030)
Tissue softening accompanies the ripening of many fruit and initiates the processes of irreversible deterioration. Expansins are plant cell wall proteins proposed to disrupt hydrogen bonds within the cell wall polymer matrix. Expression of specific expansin genes has been observed in tomato (Lycopersicon esculentum) meristems, expanding tissues, and ripening fruit. It has been proposed that a tomato ripening-regulated expansin might contribute to cell wall polymer disassembly and fruit softening by increasing the accessibility of specific cell wall polymers to hydrolase action. To assess whether ripening-regulated expansins are present in all ripening fruit, we examined expansin gene expression in strawberry (Fragaria x ananassa Duch.). Strawberry differs significantly from tomato in that the fruit is derived from receptacle rather than ovary tissue and strawberry is non-climacteric. A full-length cDNA encoding a ripening-regulated expansin, FaExp2, was isolated from strawberry fruit. The deduced amino acid sequence of FaExp2 is most closely related to an expansin expressed in early tomato development and to expansins expressed in apricot fruit rather than the previously identified tomato ripening-regulated expansin, LeExp1. Nearly all previously identified ripening-regulated genes in strawberry are negatively regulated by auxin. Surprisingly, FaExp2 expression was largely unaffected by auxin. Overall, our results suggest that expansins are a common component of ripening and that non-climacteric signals other than auxin may coordinate the onset of ripening in strawberry. (+info)
Arginine 197 of the cholecystokinin-A receptor binding site interacts with the sulfate of the peptide agonist cholecystokinin.
(36/2030)
The knowledge of the binding sites of G protein-coupled cholecystokinin receptors represents important insights that may serve to understand their activation processes and to design or optimize ligands. Our aim was to identify the amino acid of the cholecystokinin-A receptor (CCK-AR) binding site in an interaction with the sulfate of CCK, which is crucial for CCK binding and activity. A three-dimensional model of the [CCK-AR-CCK] complex was built. In this model, Arg197 was the best candidate residue for a ionic interaction with the sulfate of CCK. Arg197 was exchanged for a methionine by site-directed mutagenesis. Wild-type and mutated CCK-AR were transiently expressed in COS-7 cells for pharmacological and functional analysis. The mutated receptor on Arg197 did not bind the agonist radioligand 125I-BH-[Thr, Nle]-CCK-9; however, it bound the nonpeptide antagonist [3H]-SR27,897 as the wild-type receptor. The mutant was approximately 1,470- and 3,200-fold less potent than the wild-type CCK-AR to activate G proteins and to induce inositol phosphate production, respectively. This is consistent with the 500-fold lower potency and 800-fold lower affinity of nonsulfated CCK relative to sulfated CCK on the wild-type receptor. These data, together with those showing that the mutated receptor failed to discriminate nonsulfated and sulfated CCK while it retained other pharmacological features of the CCK-AR, strongly support an interaction between Arg197 of the CCK-AR binding site and the sulfate of CCK. In addition, the mutated CCK-AR resembled the low affinity state of the wild-type CCK-AR, suggesting that Arg197-sulfate interaction regulates conformational changes of the CCK-AR that are required for its physiological activation. (+info)
Molecular and biochemical evidence for the involvement of calcium/calmodulin in auxin action.
(37/2030)
The use of (35)S-labeled calmodulin (CaM) to screen a corn root cDNA expression library has led to the isolation of a CaM-binding protein, encoded by a cDNA with sequence similarity to small auxin up RNAs (SAURs), a class of early auxin-responsive genes. The cDNA designated as ZmSAUR1 (Zea mays SAURs) was expressed in Escherichia coli, and the recombinant protein was purified by CaM affinity chromatography. The CaM binding assay revealed that the recombinant protein binds to CaM in a calcium-dependent manner. Deletion analysis revealed that the CaM binding site was located at the NH(2)-terminal domain. A synthetic peptide of amino acids 20-45, corresponding to the potential CaM binding region, was used for calcium-dependent mobility shift assays. The synthetic peptide formed a stable complex with CaM only in the presence of calcium. The CaM affinity assay indicated that ZmSAUR1 binds to CaM with high affinity (K(d) approximately 15 nM) in a calcium-dependent manner. Comparison of the NH(2)-terminal portions of all of the characterized SAURs revealed that they all contain a stretch of the basic alpha-amphiphilic helix similar to the CaM binding region of ZmSAUR1. CaM binds to the two synthetic peptides from the NH(2)-terminal regions of Arabidopsis SAUR-AC1 and soybean 10A5, suggesting that this is a general phenomenon for all SAURs. Northern analysis was carried out using the total RNA isolated from auxin-treated corn coleoptile segments. ZmSAUR1 gene expression began within 10 min, increased rapidly between 10 and 60 min, and peaked around 60 min after 10 microM alpha-naphthaleneacetic acid treatment. These results indicate that ZmSAUR1 is an early auxin-responsive gene. The CaM antagonist N-(6-aminohexyl)5-chloro-1-naphthalenesulfonamide hydrochloride inhibited the auxin-induced cell elongation but not the auxin-induced expression of ZmSAUR1. This suggests that calcium/CaM do not regulate ZmSAUR1 at the transcriptional level. CaM binding to ZmSAUR1 in a calcium-dependent manner suggests that calcium/CaM regulate ZmSAUR1 at the post-translational level. Our data provide the first direct evidence for the involvement of calcium/CaM-mediated signaling in auxin-mediated signal transduction. (+info)
The axr6 mutants of Arabidopsis thaliana define a gene involved in auxin response and early development.
(38/2030)
The indolic compound auxin regulates virtually every aspect of plant growth and development, but its role in embryogenesis and its molecular mechanism of action are not understood. We describe two mutants of Arabidopsis that define a novel gene called AUXIN-RESISTANT6 (AXR6) which maps to chromosome 4. Embryonic development of the homozygous axr6 mutants is disrupted by aberrant patterns of cell division, leading to defects in the cells of the suspensor, root and hypocotyl precursors, and provasculature. The homozygous axr6 mutants arrest growth soon after germination lacking a root and hypocotyl and with severe vascular pattern defects in their cotyledons. Whereas previously described mutants with similar developmental defects are completely recessive, axr6 heterozygotes display a variety of morphological and physiological alterations that are most consistent with a defect in auxin physiology or response. The AXR6 gene is likely to be important for auxin response throughout the plant, including early development. (+info)
Expression of a pathogen-induced gene can be mimicked by auxin insensitivity.
(39/2030)
Following perception of a pathogenic attack, plants are able to develop a strong response with the corresponding activation of a plethora of defense-related genes. In this study we have characterized the mode of expression of the CEVI-1 gene from tomato plants, which encodes an anionic peroxidase. CEVI-1 expression is induced during the course of compatible viral and subviral infections, like many other defense-related genes, but is induced neither in incompatible interactions nor by signal molecules such as salicylic acid, ethylene, or methyl jasmonate. Additionally, CEVI-1 is induced in detached leaf tissues following a pathway distinct from that related to the classical wound response. We also describe the characterization of the structural CEVI-1 gene and compare the mode of expression in different transgenic plant species harboring a CEVI-1::GUS construct. Furthermore, we have isolated mutants in Arabidopsis, called dth mutants, that are deregulated in the control of expression of this gene. From the initial analysis of some of these mutants it seems that activation of CEVI-1 gene expression correlates with a defect in the perception of auxins by the plant. All these results may suggest that, during systemic infections with viruses, auxin homeostasis is one of the components participating in the regulation of the overall defense response. (+info)
Expression profiles of 22 novel molecular markers for organogenetic pathways acting in alfalfa nodule development.
(40/2030)
During symbiotic nodule development, a variety of molecular signals of rhizobia and plant origin are likely to be involved in the control of the expression of specific genes in the legume Medicago sativa (alfalfa). Twenty-two new, nodule-associated Expressed Sequence Tags (ESTs, MsNod clones) as well as 16 clones for previously reported alfalfa nodulins were identified by cold-plaque screening. Protein homologs were found for 10 of the 22 MsNod-encoded polypeptides, revealing putative novel functions associated with this symbiosis. Expression of these MsNod genes was investigated in spontaneous nodules (generated in the absence of bacteria), in nodules induced by a Sinorhizobium meliloti wild-type strain and Eps- and Bac- mutant derivatives, as well as in roots inoculated with a Nod- mutant strain. This analysis enabled us to correlate plant gene expression with the different stages of nodule ontogeny and invasion. The effect of phytohormones on MsNod gene expression was analyzed in cytokinin- and auxin-treated alfalfa roots. Cytokinin induced the accumulation of seven MsNod transcripts, four of them were also regulated by the synthetic auxin 2,4-D (2,4-dichlorophenoxyacetic acid). Comparison of MsNod expression profiles in wild-type and transgenic M. truncatula roots overexpressing the early nodulin Enod40 suggested that one clone, the M. sativa L3 ribosomal protein homolog (MsNod377), is a putative component of an Enod40-dependent pathway acting during nodule development. These novel molecular markers may help in the investigation of gene networks and regulatory circuits controlling nodule organogenesis. (+info)