RESUMEN
Pectin in the primary plant cell wall is thought to be responsible for its porosity, charge density, and microfibril spacing and is the main component of the middle lamella. Plant-parasitic nematodes secrete cell wall-degrading enzymes that macerate the plant tissue, facilitating the penetration and migration within the roots. In sedentary endoparasitic nematodes, these enzymes are released only during the migration of infective juveniles through the root. Later, nematodes manipulate the expression of host plant genes, including various cell wall enzymes, in order to induce specific feeding sites. In this study, we investigated expression of two Arabidopsis pectate lyase-like genes (PLL), PLL18 (At3g27400) and PLL19 (At4g24780), together with pectic epitopes with different degrees of methylesterification in both syncytia induced by the cyst nematode Heterodera schachtii and giant cells induced by the root-knot nematode Meloidogyne incognita. We confirmed upregulation of PLL18 and PLL19 in both types of feeding sites with quantitative reverse-transcriptase polymerase chain reaction (RT-PCR) and in situ RT-PCR. Furthermore, the functional analysis of mutants demonstrated the important role of both PLL genes in the development and maintenance of syncytia but not giant cells. Our results show that both enzymes play distinct roles in different infected root tissues as well as during parasitism of different nematodes.
Asunto(s)
Arabidopsis/enzimología , Interacciones Huésped-Parásitos , Enfermedades de las Plantas/parasitología , Polisacárido Liasas/metabolismo , Tylenchida/fisiología , Tylenchoidea/fisiología , Animales , Arabidopsis/citología , Arabidopsis/genética , Arabidopsis/parasitología , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Pared Celular/metabolismo , Femenino , Regulación de la Expresión Génica de las Plantas , Genes Reporteros , Células Gigantes/citología , Masculino , Mutagénesis Insercional , Fenotipo , Raíces de Plantas/citología , Raíces de Plantas/enzimología , Raíces de Plantas/genética , Raíces de Plantas/parasitología , Brotes de la Planta/citología , Brotes de la Planta/enzimología , Brotes de la Planta/genética , Brotes de la Planta/parasitología , Polisacárido Liasas/genética , Plantones/citología , Plantones/enzimología , Plantones/genética , Plantones/parasitología , Especificidad de la Especie , Tylenchida/citología , Tylenchoidea/citología , Regulación hacia ArribaRESUMEN
Thionins are a group of antimicrobial polypeptides that form part of the plant's defense mechanism against pathogens. The Thi 2.1 thionin gene of Arabidopsis thaliana has been shown to be inducible by jasmonic acid (JA), an oxylipin-like hormone derived from oxygenated linolenic acid and synthesized via the octadecanoid pathway. The JA-dependent regulation of the Thi 2.1 gene has been exploited for setting up a genetic screen for the isolation of signal transduction mutants that constitutively express the Thi 2.1 gene. Ten cet-mutants have been isolated which showed a constitutive expression of the thionin gene. Allelism tests revealed that they represent at least five different loci. Some mutants are dominant, others recessive, but all cet mutations behaved as monogenic traits when backcrossed with Thi 2.1-GUS plants. Some of the mutants overproduce JA and its bioactive precursor 12-oxophytodienoic acid (OPDA) up to 40-fold while others have the same low levels as the control wildtype plants. Two of the mutants showed a strong induction of both the salicylic acid (SA)- and the JA-dependent signaling pathways, while the majority seems to be affected only in the octadecanoid pathway. The Thi 2.1 thionin gene and the Pdf 1.2 defensin gene are activated independently, though both are regulated by JA. The cet-mutants, except for one, also show a spontaneous leaf cell necrosis, a reaction often associated with the systemic acquired resistance (SAR) pathway.
Asunto(s)
Péptidos Catiónicos Antimicrobianos , Proteínas de Arabidopsis , Arabidopsis/microbiología , Ciclopentanos/metabolismo , Ácidos Grasos Insaturados/metabolismo , Mutación , Enfermedades de las Plantas/genética , Reguladores del Crecimiento de las Plantas/metabolismo , Ácidos Esteáricos/metabolismo , Arabidopsis/genética , Segregación Cromosómica , Cruzamientos Genéticos , Fusarium , Lectinas/biosíntesis , Oxilipinas , Lectinas de Plantas , Proteínas de Plantas/biosíntesis , Ácido Salicílico/metabolismo , Transducción de Señal/genéticaRESUMEN
In seedlings of Arabidopsis thaliana the thionin gene Thi2.1 is inducible by methyl jasmonate, wounding, silver nitrate, coronatine, and sorbitol. We have used a biochemical and genetic approach to test the signal transduction of these different inducers. Both exogenously applied jasmonates and jasmonates produced endogenously upon stress induction, lead to GUS expression in a Thi2.1 promoter-uidA transgenic line. No GUS expression was observed in a coil mutant background which lacks jasmonate perception whereas methyl jasmonate and coronatine but not the other inducers were able to overcome the block in jasmonic acid production in a fad3-2 fad7-2 fad8 mutant background. Our results show conclusively that all these inducers regulate Thi2-1 gene expression via the octadecanoid pathway.
Asunto(s)
Antifúngicos , Proteínas de Arabidopsis , Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Proteínas de Plantas/biosíntesis , Proteínas de Plantas/genética , Ácidos Esteáricos/metabolismo , Péptidos Catiónicos Antimicrobianos , Arabidopsis/efectos de los fármacos , Arabidopsis/metabolismo , Ciclopentanos/antagonistas & inhibidores , Ciclopentanos/metabolismo , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Mutación , Oxilipinas , Proteínas de Plantas/metabolismo , Nitrato de Plata/farmacología , Sorbitol/farmacologíaRESUMEN
The Arabidopsis thaliana Thi2.1 gene is inducible by necrotrophic fungi through a signal transduction pathway different from that for pathogenesis-related (PR) proteins. We have identified three ecotypes that are susceptible (Col-2, Ler, and Ws) and two ecotypes that are resistant (Mt-0 and Uk-4) to spray inoculation with Fusarium oxysporum f. sp. matthiolae. The Thi2.1 transcript level after infection correlates with resistance, being 5 to 10 times higher in the resistant than in the susceptible ecotypes. The beta-glucuronidase (GUS) expression of a Thi2.1-promoter-uidA fusion (with a promoter derived from Col-2) is on the average almost 10 times higher in the Uk-4 background than in the Col-2 background. This confirms the results obtained by Northern (RNA) blots and indicates that Uk-4, and probably other resistant ecotypes too, might have a more sensitive recognition system for F. oxysporum f. sp. matthiolae or might have a signal transduction system that gives a higher amplification of the original recognition signal. Our results suggest a role of the Thi2.1 gene in resistance against F. oxysporum f. sp. matthiolae and perhaps other necrotrophic fungi.
Asunto(s)
Arabidopsis/genética , Fusarium/fisiología , Regulación Fúngica de la Expresión Génica/fisiología , Regulación de la Expresión Génica de las Plantas/fisiología , Genes de Plantas , Arabidopsis/enzimología , Arabidopsis/microbiología , Fusarium/crecimiento & desarrollo , Glucuronidasa/genética , Plantas Modificadas GenéticamenteRESUMEN
The Arabidopsis Thi2.1 thionin gene was cloned and sequenced. The promoter was fused to the uidA gene and stably transformed into Arabidopsis to study its regulation. GUS expression levels correlated with the steady-state levels of Thi2.1 mRNA, thus demonstrating that the promoter is sufficient for the regulation of the Thi2.1 gene. The sensitivity of the Thi2.1 gene to methyl jasmonate was found to be developmentally determined. Systemic and local expression could be induced by wounding and inoculation with Fusarium oxysporum f sp. matthiolae. A deletion analysis of the promoter identified a fragment of 325 bp upstream of the start codon, which appears to contain all the elements necessary for the regulation of the Thi2.1 gene. These results support the view that thionins are defence proteins, and indicate the possibility that resistance of Arabidopsis plants to necrotrophic fungal pathogens is mediated through the octadecanoid pathway.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Proteínas de Plantas/genética , Agrobacterium tumefaciens/patogenicidad , Péptidos Catiónicos Antimicrobianos , Arabidopsis/microbiología , Secuencia de Bases , Clonación Molecular , ADN de Plantas , Regulación del Desarrollo de la Expresión Génica , Glucuronidasa/genética , Datos de Secuencia Molecular , Plantas Modificadas Genéticamente , Regiones Promotoras Genéticas , Eliminación de SecuenciaRESUMEN
Viscotoxins are a group of toxic thionins found in several mistletoe species. The constitutive CaMV-omega promoter was used to drive the expression of the viscotoxin A3 cDNA from Viscum album in transgenic Arabidopsis thaliana C24. Lines with high viscotoxin A3 levels in all parts of the plant were selected and tested for resistance against the clubroot pathogen Plasmodiophora brassicae. The transgenic lines were more resistant to infection by this pathogen than the parental line.
Asunto(s)
Arabidopsis/genética , Arabidopsis/parasitología , Eucariontes/patogenicidad , Muérdago/genética , Preparaciones de Plantas , Proteínas de Plantas , Plantas Medicinales , Toxinas Biológicas/genética , Animales , Secuencia de Bases , Cartilla de ADN/genética , ADN de Plantas/genética , Expresión Génica , Genes de Plantas , Enfermedades de las Plantas/parasitología , Plantas Modificadas Genéticamente , Proteínas Inactivadoras de Ribosomas Tipo 2RESUMEN
Thionins are antimicrobial proteins that are thought to be involved in plant defense. Concordant with this view, we have recently shown that the Arabidopsis thionin Thi2.1 gene is inducible by phytopathogenic fungi. Here, we demonstrate that constitutive overexpression of this thionin enhances the resistance of the susceptible ecotype Columbia (Col-2) against attack by Fusarium oxysporum f sp matthiolae. Transgenic lines had a reduced loss of chlorophyll after inoculation and supported significantly less fungal growth on the cotyledons, as evaluated by trypan blue staining. Moreover, fungi on cotyledons of transgenic lines had more hyphae with growth anomalies, including hyperbranching, than on cotyledons of the parental line. No transcripts for pathogenesis-related PR-1, PR-5, or the pathogen-inducible plant defensin Pdf1.2 could be detected in uninoculated transgenic seedlings, indicating that all of the observed effects of the overexpressing lines are most likely the result of the toxicity of the THI2.1 thionin. Our findings strongly support the view that thionins are defense proteins.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis/genética , Fusarium/patogenicidad , Proteínas de Plantas/metabolismo , Péptidos Catiónicos Antimicrobianos , Arabidopsis/microbiología , Clonación Molecular , Plantas Modificadas GenéticamenteRESUMEN
Plant defensins, formerly named gamma-thionins, are a group of small, cysteine-rich, basic, and antimicrobial plant proteins. Random sequencing of expressed sequence tags (ESTs) in Arabidopsis thaliana has revealed several different plant defensin genes in this plant species which can be grouped into two subfamilies. We have used one EST of each subfamily to study the expression of the corresponding genes in A. thaliana. Pdf2.3 is constitutively expressed in seedlings, rosettes, flowers, and siliques and is not inducible in seedlings either by methyl jasmonate, salicylate, ethephon, and silver nitrate or by several different phytopathogenic fungi. The expression of a second gene, Pdf1.2, is in untreated plants only detectable in rosettes. In seedlings, it is inducible by methyl jasmonate, silver nitrate, and different phytopathogenic fungi, notably Fusarium oxysporum f. sp. matthiolae. The regulation of Pdf1.2 resembles that of the pathogen-inducible thionin gene Thi2.1.
Asunto(s)
Antifúngicos/farmacología , Proteínas de Arabidopsis , Arabidopsis/inmunología , Defensinas , Familia de Multigenes , Proteínas de Plantas/genética , Lugares Marcados de Secuencia , Alternaria/inmunología , Secuencia de Aminoácidos , Péptidos Catiónicos Antimicrobianos , Arabidopsis/genética , Fusarium/inmunología , Expresión Génica , Datos de Secuencia Molecular , Proteínas de Plantas/químicaRESUMEN
We compared the organ specificity and the strength of different constitutive (CaMV-35S, CaMV-35Somega, Arabidopsis ubiquitin UBQ1, and barley leaf thionin BTH6 promoter) and one inducible promoter (soybean heat-shock promoter Gmhsp17.3) in stably transformed Arabidopsis thaliana plants. For this purpose we constructed a set of plant expression vectors equipped with the different promoters. Using the uidA reporter gene we could show that the CaMV-35S promoter has the highest expression level which was enhanced two- to threefold by the addition of a translational enhancer (TMV omega element) without altering the organ specificity of the promoter. The barley leaf thionin promoter was almost inactive in the majority of lines whereas the ubiquitin promoter exhibited an intermediate strength. The heat-shock promoter was inducible up to 18-fold but absolute levels were lower than in the case of the ubiquitin promoter. Conclusive quantitative results for different organs and developmental stages were obtained by the analysis of 24 stably transformed lines per promoter construct.
Asunto(s)
Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Vectores Genéticos , Plantas Modificadas Genéticamente , Regiones Promotoras Genéticas , Péptidos Catiónicos Antimicrobianos , Secuencia de Bases , Glucuronidasa/genética , Proteínas de Choque Térmico/genética , Respuesta al Choque Térmico , Datos de Secuencia Molecular , Hojas de la Planta/metabolismo , Proteínas de Plantas/genética , Raíces de Plantas/metabolismo , Semillas/crecimiento & desarrollo , Semillas/metabolismo , Distribución Tisular , Transformación Genética , Transgenes , Ubiquitinas/genéticaRESUMEN
Two cDNAs encoding thionin preproteins have been isolated from Arabidopsis thaliana. The corresponding genes have been designated Thi2.1 and Thi2.2. Southern blot analysis suggests that A. thaliana most probably contains single genes for both thionins. Thi2.2 transcripts have a low basal level in seedlings and show circadian variation. Thi2.2 transcripts were also detected in rosette leaves. No potent elicitors have been found for the Thi2.2 gene. Transcripts of the Thi2.1 gene are not detectable in seedlings but are present in rosette leaves and at a very high level in flowers and in siliques. The expression of the Thi2.1 gene is highly inducible in seedlings by pathogens, silver nitrate, and methyl jasmonate, but not by salicylate, indicating that the gene is induced by a signal transduction pathway that is at least partly different from that for the pathogenesis-related proteins.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis/genética , Regulación de la Expresión Génica , Genes de Plantas , Proteínas de Plantas/genética , Transducción de Señal , Secuencia de Aminoácidos , Péptidos Catiónicos Antimicrobianos , Arabidopsis/microbiología , Secuencia de Bases , Northern Blotting , Southern Blotting , ADN Complementario/genética , Fusarium/patogenicidad , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Datos de Secuencia Molecular , Micosis , Proteínas de Plantas/metabolismo , Análisis de Secuencia de ADNRESUMEN
Patients history, clinical findings, pulmonary function and necessary therapy are combined for a diagram, which should easily allow to grade childhood asthma for general practitioners, pediatricians and consultants of the health insurance. The grading system is based on the cited literature, dealing in part with the matter, and on the international consensus report on asthma.
Asunto(s)
Asma/clasificación , Evaluación de la Discapacidad , Asma/fisiopatología , Niño , Testimonio de Experto , Humanos , Mediciones del Volumen PulmonarRESUMEN
Cochliobolus heterostrophus, a heterothallic Ascomycete, has a single mating type locus with two alternate forms called MAT-1 and MAT-2. MAT-1 was cloned by complementing a MAT-2 strain using a cosmid library from a MAT-1 strain and screening for a homothallic transformant. The cosmid recovered from this transformant was able to re-transform a MAT-2 strain to homothallism and MAT identity was proven by restriction fragment length polymorphism and conventional genetic mapping. All homothallic transformants could mate with either MAT-1 or MAT-2 strains, although the number of ascospores produced by self matings or crosses to MAT-2 strains was low. Progeny of selfed homothallic transformants were themselves homothallic. MAT-2 was cloned by probing a cosmid library from a MAT-2 strain with a fragment of insert DNA from a MAT-1 cosmid. A 1.5 kb subclone of either MAT-containing cosmid was sufficient to confer mating function in transformants. Examination of the DNA sequence of these subclones revealed that MAT-1 and MAT-2 contain 1297 bp and 1171 bp, respectively, of completely dissimilar DNA flanked by DNA common to both mating types. Putative introns were found (one in each MAT gene) which, when spliced out, would yield open reading frames (ORFs) that occupied approximately 90% of the dissimilar DNA sequences. Translation of the MAT-1 ORF revealed similarity to the Neurospora crassa MATA, Podospora anserina mat-, and Saccharomyces cerevisiae MAT alpha 1 proteins; translation of the MAT-2 ORF revealed similarity to the N. crassa MATa, P. anserina mat+, and Schizosaccharomyces pombe mat-Mc proteins. These gene products are all proven or proposed DNA binding proteins. Those with similarity to MAT-2 are members of the high mobility group.
Asunto(s)
Proteínas de Unión al ADN/genética , Proteínas Fúngicas , Genes Fúngicos , Genes del Tipo Sexual de los Hongos , Xylariales/genética , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , Cósmidos , Cruzamientos Genéticos , ADN de Hongos/genética , ADN de Hongos/aislamiento & purificación , Hongos/genética , Biblioteca de Genes , Vectores Genéticos , Intrones , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , Biosíntesis de Proteínas , Mapeo Restrictivo , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , Transformación GenéticaRESUMEN
A novel class of highly abundant polypeptides with antifungal activity has been detected in cell walls of barley leaves. Similar polypeptides known as thionins occur not only in monocotyledonous but also in various dictoyledonous plants. The leaf-specific thionins of barley are encoded by a complex multigene family, which consists of at least 50-100 members per haploid genome. All of these genes are confined to chromosome 6. The toxicity of these thionins for plant pathogenic fungi and the fact that their synthesis can also be triggered by pathogens strongly suggest that thionins are a naturally occurring, inducible plant protein possibly involved in the mechanism of plant defence against microbial infections.
Asunto(s)
Asma/diagnóstico , Alérgenos , Niño , Hipersensibilidad a los Alimentos/diagnóstico , Humanos , MétodosRESUMEN
The protective and curative effect of Th 1165a (Fenoterol) and Salbutamol on the allergic rhinitis is reported. 70 patients with positive nasal provocation tests (speezing, rhinitis and eosinophilia of the secretions) were tested some days later with the same allergen after application of 100 mug TH 1165a. Protective effect could be shown in 64 patients who did not react with clinical symptoms. Another 85 patients were treated in the same way after a positive nasal provocation test and the clinical symptoms disappered within 5 minutes. In 10 patients nasal resistance was measured objectively by bodyplethysmography. Salbutamol showed the same protective and curative effect in 30 patients proved in the same way as described before. Application of the substances in the contralateral nostril had no systemic effect.
Asunto(s)
Albuterol/administración & dosificación , Metaproterenol/administración & dosificación , Rinitis Alérgica Estacional/prevención & control , Resistencia de las Vías Respiratorias/efectos de los fármacos , Humanos , Fenoles/administración & dosificación , Factores de TiempoRESUMEN
Systematic measurements of blood lead levels in samples from 176 mature and premature neonates and 59 mothers in the western Ruhr-Rhine area demonstrated that at least 30% of statistical variance in blood lead levels depended on lead immission at the mother's place of residence. The close correlation between neonatal and maternal blood lead levels was confirmed. Identical and non-identical twins had almost the same blood lead levels. Using a biological system for determining lead immission, blood lead levels in areas not covered by the present study can be prognosticated.