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P Vera

Publications and source records attributed to P Vera.

At least 19 recordsLinked to original sources
PubMed2000

Expression of a pathogen-induced gene can be mimicked by auxin insensitivity.

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.

Arabidopsis↗
PubMed2000

Characterization of P69E and P69F, two differentially regulated genes encoding new members of the subtilisin-like proteinase family from tomato plants.

Subtilisin-like proteins represent an ancient family of serine proteases that are extremely widespread in living organisms. We report here the structure and genomic organization of two new transcriptionally active genes encoding proteins that belong to the P69 family of subtilisin-like proteases from tomato (Lycopersicon esculentum) plants. The two new members, P69E and P69F, are organized in a cluster and arranged in a tandem form. mRNA expression analysis and studies of transgenic Arabidopsis plants transformed with promoter-beta-glucuronidase fusions for each of these two genes revealed that they are differentially regulated, with each showing a highly specific mRNA expression pattern. P69E mRNA is expressed only in roots, while P69F mRNA is expressed only in hydathodes. A comparison of all the P69 amino acid sequences, gene structure, expression profiles, and clustered organization suggests a working model for P69 gene family evolution.

Amino Acid Sequence↗
PubMed2000

Local and systemic induction of two defense-related subtilisin-like protease promoters in transgenic Arabidopsis plants. Luciferin induction of PR gene expression.

Following a pathogenic attack, plants are able to mount a defense response with the coordinated activation of a battery of defense-related genes. In this study we have characterized the mode of expression of the P69B and P69C genes from tomato (Lycopersicon esculentum Mill.), which encodes two closely related subtilisin-like proteases associated with the defense response. We have compared the mode of gene regulation in heterologous transgenic Arabidopsis plants harboring promoter-beta-glucuronidase (GUS) and promoter-luciferase (LUC) gene fusions for these two genes. These studies revealed that the P69B and P69C promoters are induced by salicylic acid as well as during the course of both a compatible and an incompatible interaction with Pseudomonas syringae. Furthermore, P69B and P69C expression takes place in both the local and the distal (noninoculated) leaves upon inoculation with bacteria but following different and unique tissue-specific patterns of expression that are also different to that described for most other classical PR genes. Also, we report that luciferin, the substrate for the reporter luciferase (LUC) gene, is able to activate expression of PR genes, and this may pose a problem when using this gene reporter system in studies related to plant defense.

Arabidopsis↗
PubMed2000

Arabidopsis dth9 mutation identifies a gene involved in regulating disease susceptibility without affecting salicylic acid-dependent responses.

To determine which components of the plant defense response make important contributions to limiting pathogen attack, an M(2) mutagenized population of a transgenic Arabidopsis line was screened for mutants showing constitutive expression of beta-glucuronidase activity driven by the promoter region of the CEVI-1 gene. The CEVI-1 gene originally was isolated from tomato plants and has been shown to be induced in susceptible varieties of tomato plants by virus infection in a salicylic acid-independent manner. We report here the characterization of a recessive mutant, detachment9 (dth9). This mutant is more susceptible to both virulent and avirulent forms of the oomycete Peronospora and also exhibits increased susceptibility to the moderately virulent bacterial pathogen Pseudomonas syringae pv maculicola ES4326. However, this mutant is not affected in salicylic acid metabolism and shows normal expression of pathogenesis-related (PR) genes after pathogen attack. Furthermore, after inoculation with avirulent pathogens, the dth9 mutant shows a compromised systemic acquired resistance response that cannot be complemented by exogenous application of salicylic acid, although this molecule is able to promote normal activation of PR genes. Therefore, the dth9 mutation defines a regulator of disease susceptibility that operates upstream or independently of salicylic acid. Pleiotropy is also evident in the dth9 mutant in the sense that the shoots of dth9 plants are insensitive to the exogenously applied auxin analog 2,4-dichlorophenoxyacetic acid.

Arabidopsis↗
PubMed2000

[Primary intraosseous rhabdomyosarcoma].

We report a case of primary intraosseous pleiomorphic rhabdomyosarcoma located in the pelvis of a 21-year-old woman followed for 4 years. The lytic tumor involved the acetabulum and the isthma with moderate extension to soft tissue. First line chemotherapy was unable to arrest tumor progression. Hemipelvectomy with saddle prosthesis reconstruction was performed, but septic complications dictated a secondary inter-ilio-abdominal amputation. Recurrence-free remission was achieved for 4 years, suggesting this was indeed a primary tumor. Primary intraosseous rhabdomyosarcomas are exceptional. Bone localizations generally suggest metastasis from a primary tumor often situated in an intraperitoneal localization. When search for a primary tumor is negative, intraosseous lesions can be considered as primary tumors warranting curative treatment. Radical surgical resection is recommended within the framework of a multidiscipinary management protocol associating radiotherapy and chemotherapy to improve prognosis.

Adult↗
PubMed1999

A genomic cluster containing four differentially regulated subtilisin-like processing protease genes is in tomato plants.

Screening of a genomic library from tomato plants (Lycopersicon esculentum) with a cDNA probe encoding a subtilisin-like protease (PR-P69) that is induced at the transcriptional level following pathogen attack (Tornero, P., Conejero, V., and Vera, P. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 6332-6337) resulted in the isolation of a cluster of genomic clones that comprise a tandem of four different subtilisin-like protease genes (P69A, P69B, P69C, and P69D). Sequence analyses and comparison of the encoded proteins revealed that all are closely related (79 to 88% identity), suggesting that all are derived from a common ancestral gene. mRNA expression analysis as well as studies of transgenic plants transformed with promoter-beta-glucuronidase fusions for each of these genes revealed that the four genes exhibit differential transcriptional regulation and expression patterns. P69A and P69D are expressed constitutively, but with different expression profiles during development, whereas the P69B and P69C genes show expression following infection with Pseudomonas syringae and are also up-regulated by salicylic acid. We propose that these four P69-like proteases, as members of a complex gene family of plant subtilisin-like proteases, may be involved in a number of specific proteolytic events that occur in the plant during development and/or pathogenesis.

Amino Acid Sequence↗
PubMed1999

Developmental regulation of a cytosolic ascorbate peroxidase gene from tomato plants.

This report describes the characterisation of a gene (APX20) from tomato plants that encodes a cytosolic isoform of ascorbate peroxidase which is involved in the detoxification of intracellular H2O2. Expression analysis of promoter-GUS fusions in transgenic plants reveals that the gene is under strict developmental control, becoming transcriptionally active in cells which are apparently undergoing mechanical stimulation generated during the different phases of growth. We show that the APX20 gene contains a large 5' leader intron which is required to confer constitutive gene expression in leaves, but not in other organs of the plant. Based on these observations, we propose that the observed transcriptional regulation of this gene may constitute a basic mechanism for deployment of antioxidative defences in plants, which act to limit the deleterious effects of H2O2 generated during normal plant development.

5' Untranslated Regions↗
PubMed1999

Identification of a novel peptide motif that mediates cross-linking of proteins to cell walls.

A cDNA clone representing a member of a novel class of cell wall proteins was isolated from tobacco plants. We have designated this protein NtTLRP for tyrosine- and lysine-rich protein. It is structurally related to the previously identified TLRP from tomato plants, sharing a high amino-acid sequence similarity at the C-terminal region. This region contains what appears to be a novel peptide motif which we call CD for cysteine-rich domain, and which is common to several other cell-wall proteins. By using a functional test in transgenic plants, we demonstrate that the presence of the CD domain is per se sufficient to cross-link previously soluble proteins to the cell wall. We present evidence that NtTLRP is cross-linked and specifically localizes to the cell wall of lignified cells. The highly localized deposition of NtTLRP in these cells indicates that this class of cell-wall proteins may have a specialized function in the formation of xylem tissue.

Amino Acid Sequence↗
PubMed1999

A tomato homeobox gene (HD-zip) is involved in limiting the spread of programmed cell death.

Antisense suppression in transgenic tomato plants of H52, a gene encoding a new homeodomain protein of the HD-Zip class, produces a conditional lethal phenotype. The transgenic lines that survive exhibit spontaneous misregulation of cell death control in leaves, which, once initiated, propagates and engulfs the entire leaf. Activation of defence genes, over-accumulation of ethylene and conjugated salicylic acid, and growth reduction of virulent pathogens also occurs in these plants. In wild-type plants, H52 is up-regulated upon infection, mirroring the generation of the oxidative burst which normally precedes the hypersensitive response (HR). Thus, H52 appears to be a transcription factor involved in cellular protection by limiting spread of programmed cell death in plants.

Amino Acid Sequence↗
PubMed1997

Identification of a new pathogen-induced member of the subtilisin-like processing protease family from plants.

By using biochemical, immunological, and molecular strategies we have identified and cloned a cDNA encoding a protease from tomato (Lycopersicon esculentum) plants (P69B) that is part of a proteolytic system activated in the plant as a result of infection with citrus exocortis viroid. This new protease is closely related, in terms of amino acid sequence and structural organization, to the previously identified pathogenesis-related subtilisin-like protease (Tornero, P., Conejero, V., and Vera, P. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 6332-6337). The 745-residue amino acid sequence of P69B begins with a cleavable signal peptide, contains a prodomain and a 631-residue mature domain which is homologous to the catalytic modules of bacterial subtilisins and eukaryotic Kex2-like proteases. Within the catalytic domain, the essential Asp, His, and Ser residues that conform the catalytic triad of this family of proteases are conserved in P69B. Northern blot and reverse transcriptase-polymerase chain reaction analysis demonstrated widespread induced expression of the 2.5-kilobase hybridizing mRNA in plant tissues as a consequence of viroid infection. We propose that P69B is a member of a complex gene family of plant Kex2/subtilisin-like proteases presumably involved in a number of specific proteolytic events activated during pathogenesis in plants and that takes place in the extracellular matrix.

Amino Acid Sequence↗
PubMed1997

Adsorption of a Corticoid on Colloidal Hematite Particles of Different Geometries

An electrokinetic study of the adsorption of a glucocorticoid drug, betamethasone disodium phosphate (BMP), on hematite (alpha-Fe2O3) particles is presented in this work. We analyze the electrophoretic mobility of both spherical and elongated hematite as a function of the concentration of BMP in the dispersion medium; it is found that, even for the lowest BMP concentration, the drug molecules adsorb onto the particles, changing their surface charge from positive to negative. When the drug concentration is increased between 10(-5) and 5 x 10(-3) M, the mobility of spherical hematite shows a monotonous increase from approximately -1.5 to approximately -3.5 &mgr;ms-1/Vcm-1; in the case of elongated particles, the mobility increases with BMP concentration up to 10(-4) M, larger concentrations of the corticoid having little effect on the electrokinetic properties of the particles. The effect of pH on the electrophoretic mobility of hematite is also investigated for two constant drug concentrations (0.05 and 0.5 mM); the isoelectric point of the particles is shifted to a pH below 3, another indication of adsorption of the negatively charged phosphate groups of BMP. These qualitative reasonings are confirmed by direct spectrophotometric determinations of the amount of BMP adsorbed on both hematite samples: spherical and ellipsoidal. The effect of pH on the adsorption density, Gamma, suggests that electrostatic interactions are not the only forces responsible for adsorption. In fact, although Gamma decreases with pH for both types of particles, it remains measurable up to pH 10, when strong electric repulsion is to be expected between hematite and phosphate groups. The effect of initial BMP concentration on Gamma is also studied. It is shown that a plateau region is reached in both samples, this indicating that their surfaces get saturated with BMP: the saturation begins when the coverage is of the order of approximately 1/8 (spherical particles) and approximately 1/40 (elongated particles) of a drug monolayer. Such difference might be due to the ellipsoidal hematite particles having a lower positive surface charge, since they are synthesized in a different dispersion medium.

Journal Article↗
PubMed1997

Complementarity of lung scintigraphy and D-dimer test in pulmonary embolism.

D-dimer assay (DDA), measuring fibrin degradation products, was compared with lung scintigraphy (LS) in a prospective unselected series of 83 consecutive patients referred owing to suspicion of pulmonary embolism (PE). This patient series was also used to compare several methods of performing and interpreting LS images. The final diagnosis was established independently by a separate panel with all available information except for the result of DDA. D-dimer was determined by ELISA (threshold value 500 ng/ml). LS, including perfusion (.Q) and pseudo-ventilation (Technegas) (.V), was classified according to PIOPED, (1) immediately by the physician on duty, and (2) retrospectively by a blinded panel. A positive (19) or negative (61) diagnosis of PE was achieved in 80 patients, the prevalence of PE being 24%. Only one false-negative was noted on DDA (sensitivity=95%) but there were 42 false-positives (specificity=31%), resulting in a positive predictive value of 30% and a negative predictive value of 95%. Emergency and retrospective interpretations of LS were close (kappa=0.4). In a minority of patients, PE may be excluded with reasonable certainty if DDA is normal, resulting in a significant saving in terms of time and money.

Enzyme-Linked Immunosorbent Assay↗
PubMed1997

Two PR-1 genes from tomato are differentially regulated and reveal a novel mode of expression for a pathogenesis-related gene during the hypersensitive response and development.

Pathogenesis-related (PR) proteins form a heterogeneous family of plant proteins that are likely to be involved in defense and are inducible by pathogen attacks. One group of PRs, represented by the subfamily PR-1, are low-molecular-weight proteins of unknown biochemical function. Here we describe the cloning and characterization of two closely related genes encoding a basic and an acidic PR-1 protein (PR1b1 and PR1a2) from tomato (Lycopersicon esculentum). We present a comparative study of the mode of transcriptional regulation of these two genes in transgenic tobacco plants using a series of promoter-GUS fusions. Unexpectedly, the chimeric PR1a2/GUS gene is not induced by pathogenic signals but instead shows constitutive expression with a reproducible developmental expression pattern. It is expressed in shoot meristems, trichomes, and cortical cells as well as in vascular and nearby tissues of the mature stem. This constitutive expression pattern may represent preemption of plant defenses against potential pathogens. Conversely, the chimeric PR1b1/GUS gene does not show any constitutive expression in the plant, but it is transcriptionally activated following pathogen attack. Upon infection by tobacco mosaic virus, the PR1b1 gene is strongly activated locally in tissues undergoing the hypersensitive response but not systemically in uninoculated tissues. Furthermore, its expression is induced by both salicylic acid and ethylene precursors, two signals that coexist and apparently mediate the activation of local defenses during the hypersensitive response. We speculate that the different mode of expression of the two genes presented here, together with that reported previously for the induction of other PR-1 genes in systemic, uninoculated tissues, may all be complementary and necessary for the plant to acquire an efficient refractory state to resist pathogen attacks.

Amino Acid Sequence↗
PubMed1996

Primary structure and expression of a pathogen-induced protease (PR-P69) in tomato plants: Similarity of functional domains to subtilisin-like endoproteases.

A 69-kDa proteinase (P69), a member of the pathogenesis-related proteins, is induced and accumulates in tomato (Lycopersicon esculentum) plants as a consequence of pathogen attack. We have used the polymerase chain reaction to identify and clone a cDNA from tomato plants that represent the pathogenesis-related P69 proteinase. The nucleotide sequence analysis revealed that P69 is synthesized in a preproenzyme form, a 745-amino acid polypeptide with a 22-amino acid signal peptide, a 92-amino acid propolypeptide, and a 631-amino acid mature polypeptide. Within the mature region the most salient feature was the presence of domains homologous to the subtilisin serine protease family. The amino acid sequences surrounding Asp-146, His-203, and Ser-532 of P69 are closely related to the catalytic sites (catalytic triad) of the subtilisin-like proteases. Northern blot analysis revealed that the 2.4-kb P69 mRNA accumulates abundantly in leaves and stem tissues from viroid-infected plants, whereas the mRNA levels in tissues from healthy plants were undetectable. Our results indicate that P69, a secreted calcium-activated endopeptidase, is a plant pathogenesis-related subtilisin-like proteinase that may collaborate with other defensive proteins in a general mechanism of active defense against attacking pathogens.

Amino Acid Sequence↗
PubMed1996

Adsorption of betamethasone disodium phosphate on ethylcellulose latex: an electrokinetic study.

The use of polymer latexes as vehicles for drug transport and controlled delivery in the body is a very active research field in applied colloid science. In this work, the polymer chosen is a commercially available ethylcellulose-based material, Aquacoat. We report electrokinetic (electrophoresis) data on this polymer in the presence of the soluble glucocorticoid betamethasone disodium phosphate (BMP). The electrophoretic mobility of the particles becomes more negative as the concentration of BMP in the medium is increased between 10(-5) and 10(-3) M, this demonstrating that the negative species produced upon dissolution of BMP adsorb on the particles; this trend is reversed for higher concentrations due to the phenomenon of double layer thickness. When pH is increased at constant BMP concentration, the negative mobility increases in absolute value, but it reaches lower values than attained in the absence of BMP. This is interpreted as another consequence of adsorption, since the ionization constants of the phosphate and sulfate groups are different. Such adsorption is confirmed and measured by spectrophotometric analysis of the supernatants after centrifugation of the suspensions; it is found that the adsorption density increases with the initial concentration of BMP in the medium, an almost vertical isotherm being obtained for concentrations above 5 x 10(-3) M, possibly due to bidimensional aggregation of the adsorbed molecules and multilayer adsorption at higher concentrations. It is found that the adsorbed amount increases with the ionic strength of the medium as a consequence of screening of the particle and BMP charges, whereas these electrostatic contributions to adsorption manifest also in the pH dependence of adsorption. IR analysis of the particles after equilibration with BMP solutions of increasing concentration agrees well with adsorption data.

Adsorption↗
PubMed1996

Characterization of LRP, a leucine-rich repeat (LRR) protein from tomato plants that is processed during pathogenesis.

This paper describes the isolation and characterization of LRP, a new gene from tomato plants. The deduced amino acid sequence showed that the encoded protein is enriched in leucine, and contains interesting structural motifs. LRP contains four tandem repeats of a canonical 24 amino acid leucine-rich repeat (LRR) sequence present in different proteins that mediates molecular recognition and/or interaction processes. Genomic organization and intron-exon arrangement of LRP favor the hypothesis that the LRR domains present in LRP evolved by exon duplication and shuffling. LRP expression analysis and immunohistochemical localization studies of the encoded protein indicate that the gene is under developmental regulation exhibiting tissue-specificity, particularly in certain cell types of the stele, like phloem fibers, parenchyma cells of the protoxylem, and in the cell files that constitute the rays of the secondary xylem. It is shown that this gene is upregulated in diseased tomato plants infected with citrus exocortis viroid. However, in this pathogenic context, LRP is processed proteolytically to a lower molecular weight form by a host-induced extracellular protease. The structural characteristics of LRP, its spatio-temporal pattern of expression, and its post-translational processing during pathogenesis, suggest this protein as a candidate molecule that may mediate recognition and interaction events taking place in the plant extracellular matrix under normal and/or pathogenesis-related conditions.

Amino Acid Sequence↗
PubMed1996

Phloem-specific expression of a plant homeobox gene during secondary phases of vascular development.

This paper reports the isolation and characterization of a homeobox gene (VAHOX1) from Lycopersicon esculentum encoding a homeodomain protein that contains a leucine zipper motif. The bipartite homeodomain-leucine zipper (HD-Zip) motif has only been found in homeobox genes from dicotyledonous plants, indicating that this type of transcription factor regulates particular developmental processes in these plant species. Here, the genomic organization, sequence comparison and expression analysis of VAHOX1 are described. Transcriptional fusion of the 5' promoter region of VAHOX1 with the reporter GUS gene (VAHOX1-GUS) and expression analysis of this construct in transgenic plants indicates that VAHOX1 is specifically expressed in the phloem during phases of secondary growth. Unlike other plant homeobox genes, VAHOX1 does not appear to be expressed in meristems, and the function of VAHOX1 is considered a likely candidate molecule that may participate in the regulation of the identity and/or activity of phloem tissues during secondary phases of vascular development.

Amino Acid Sequence↗