Temperature-mediated biosynthesis of the phytotoxin phaseolotoxin by Pseudomonas syringae pv. phaseolicola depends on the autoregulated expression of the phtABC genes

Abstract Pseudomonas syringae pv. phaseolicola produces phaseolotoxin in a temperature dependent manner, being optimally synthesized between 18˚C and 20˚C, while no detectable amounts are present above 28˚C. The Pht cluster, involved in the biosynthesis of phaseolotoxin, contains 23 genes that are organized in five transcriptional units. The function of most of the genes from the Pht cluster is still unknown and little information about the regulatory circuitry leading to expression of these genes has been reported. The purpose of the present study was to investigate the participation of pht genes in the regulation of the operons coded into the Pht cluster. We conducted Northern blot, uidA fusions and reverse transcription-PCR assays of pht genes in several mutants unable to produce phaseolotoxin. This allowed us to determine that, in P. syringae pv. phaseolicola NPS3121, genes phtABC are essential to prevent their own expression at 28˚C, a temperature at which no detectable amounts of the toxin are present. We obtained evidence that the phtABC genes also participate in the regulation of the phtD, phtM and phtL operons. According to our results, we propose that PhtABC and other Pht product activities could be involved in the synthesis of the sulfodiaminophosphinyl moiety of phaseolotoxin, which indirectly could be involved in the transcriptional regulation of the phtA operon

The Pbo cluster from Pseudomonas syringae pv. phaseolicola NPS3121 is thermoregulated and required for phaseolotoxin biosynthesis

The bean (Phaseolus vulgaris) pathogen Pseudomonas syringae pv. phaseolicola NPS3121 synthe-sizes phaseolotoxin in a thermoregulated way, with optimum production at 18 °C. Gene PSPPH_4550 was previously shown to be thermoregulated and required for phaseolotoxin bio-synthesis. Here, we established that PSPPH_4550 is part of a cluster of 16 genes, the Pbo cluster, included in a genomic island with a limited distribution in P. syringae and unrelated to the posses-sion of the phaseolotoxin biosynthesis cluster. We identified typical non-ribosomal peptide syn-thetase, and polyketide synthetase domains in several of the pbo deduced products. RT-PCR and the analysis of polar mutants showed that the Pbo cluster is organized in four transcriptional units, including one monocistronic and three polycistronic. Operons pboA and pboO are both es-sential for phaseolotoxin biosynthesis, while pboK and pboJ only influence the amount of toxin produced. The three polycistronic units were transcribed at high levels at 18 °C but not at 28 °C, whereas gene pboJ was constitutively expressed. Together, our data suggest that the Pbo cluster synthesizes secondary metabolite(s), which could participate in the regulation of phaseolotoxin biosynthesis.The authors thank the Consejo Nacional de Ciencia y Tecnología (CONACyT) for the scholarship granted to Lizeth Guardado-Valdivia. The work reported was funded by research grant CB-2015-01-255155 from the CONACyT, to S. Aguilera, and by the Spanish Plan Nacional I+D+I grant AGL2017-82492-C2-2-R, from the Ministerio de Economía y Competitividad (MINECO), co-financed by the Fondo Europeo de Desarrollo Regional (FEDER), to J. Murillo

Expression of the gene for resistance to phaseolotoxin (argK) depends on the activity of genes phtABC in Pseudomonas syringae pv. Phaseolicola

Incluye material complementarioThe bacterium Pseudomonas syringae pv. phaseolicola produces phaseolotoxin in a temperature dependent manner, being optimally produced between 18 degrees C and 20 degrees C, while no detectable amounts are present above 28 degrees C. Phaseolotoxin is an effective inhibitor of ornithine carbamoyltransferase (OCTase) activity from plant, mammalian and bacterial sources and causes a phenotypic requirement for arginine. To protect the cell from its own toxin, P. syringae pv. phaseolicola synthesizes a phaseolotoxin-resistant OCTase (ROCT). The ROCT is the product of the argK gene and is synthesized only under conditions leading to phaseolotoxin synthesis. The argK gene is included in a chromosomal fragment named Pht cluster, which contains genes involved in the synthesis of phaseolotoxin. The aim of the present work was to investigate the possible involvement of other genes included in the Pht cluster in the regulation of gene argK. We conducted transcriptional analyses of argK in several mutants unable to produce phaseolotoxin, transcriptional fusions and electrophoretic mobility shift assays, which allowed us to determine that genes phtABC, located within the Pht cluster, participate in the transcriptional repression of gene argK at temperatures not permissive for phaseolotoxin biosynthesis. This repression is mediated by a protein present in both toxigenic and nontoxigenic strains of P. syringae and in E. coli, and requires the coordinated participation of phtA, phtB and phtC products in order to carry out an efficient argK repression.This work was funded by grants from CONACYT (Consejo Nacional de Ciencia y Tecnología; http://www.conacyt.mx), research grant SEP-2006-C01-49958/24089 to AAM and SA (Postdoctoral scholarship), and from the Spanish Plan Nacional I+D+I grant AGL2008-55311-CO2-01 (Ministerio de Ciencia e Innovación; http://micinn.es/), co financed by FEDER, to JM

Aplicación de un sistema de monitoreo y evaluación del perfil de egreso de las carreras Ingeniería en Informática, Administración de Empresa y Administración Agropecuaria, en el Instituto AIEP - Sede San Fernando

Tesis (Magíster en Liderazgo de Gestión y Administración Educacional)El trabajo que se presenta a continuación surge a partir de la concurrencia de tres focos específicos, la participación de los autores en el Magíster en Administración y Liderazgo Educación. Educativa, el ejercicio profesional en las áreas o módulos desarrollados, y la práctica docente de los módulos. La coincidencia mencionada se traduce en la Tesis que se expone a continuación y se juzga importante introducirla en éstos términos para que el lector pueda situarla en su contexto específico. La perspectiva interdisciplinaria a través de la cual se aborda el tema no es un detalle menor o casual, el desarrollo de la informática, la administración y la agroindustria tiene entre sus efectos más relevantes el generar fuentes de trabajo para el universo analizado. De esta forma las herramientas que se utilizan para educar a un futuro técnico o profesional en las áreas mencionadas es parte importante de su proceso creativo y de producción

Comunicación Productiva como estrategia para fortalecer la calidad en el servicio en organizaciones educativas. Caso práctico: Bachillerato 5 de Mayo de la Benemérita Universidad Autónoma de Puebla

“La evolución de las organizaciones ha ido de la mano del desarrollo social, las organizaciones educativas no han sido la excepción, sin embargo en el camino la función y objetivo de estas organizaciones se ha desvirtuado como consecuencia de un acelerado progreso influido por la tecnología y la economía de mercado. Hoy más que nunca las organizaciones educativas son corresponsables de la construcción del entramado social; muchos de los vicios y de la pérdida de visión sobre estas instituciones, tiene que ver con la economía de mercado y la globalización. Estando la organización educativa a expensas del entorno que demanda personas competentes, no competitivas, pero sin embargo los objetivos institucionales no son acordes a dichas demandas, iniciando porque los públicos estratégicos de dichas entidades tienen diversas concepciones sobre la calidad que deberían de ofertar en el servicio que brindan en el servicio educativo.

Two Homologues of the Global Regulator Csr/Rsm Redundantly Control Phaseolotoxin Biosynthesis and Virulence in the Plant Pathogen Pseudomonas amygdali pv. phaseolicola 1448A

The widely conserved Csr/Rsm (carbon storage regulator/repressor of stationary-phase metabolites) post-transcriptional regulatory system controls diverse phenotypes involved in bacterial pathogenicity and virulence. Here we show that Pseudomonas amygdali pv. phaseolicola 1448A contains seven rsm genes, four of which are chromosomal. In RNAseq analyses, only rsmE was thermoregulated, with increased expression at 18 °C, whereas the antagonistic sRNAs rsmX1, rsmX4, rsmX5 and rsmZ showed increased levels at 28 °C. Only double rsmA-rsmE mutants showed significantly altered phenotypes in functional analyses, being impaired for symptom elicitation in bean, including in planta growth, and for induction of the hypersensitive response in tobacco. Double mutants were also non-motile and were compromised for the utilization of different carbon sources. These phenotypes were accompanied by reduced mRNA levels of the type III secretion system regulatory genes hrpL and hrpA, and the flagellin gene, fliC. Biosynthesis of the phytotoxin phaseolotoxin by mutants in rsmA and rsmE was delayed, occurring only in older cultures, indicating that these rsm homologues act as inductors of toxin synthesis. Therefore, genes rsmA and rsmE act redundantly, although with a degree of specialization, to positively regulate diverse phenotypes involved in niche colonization. Additionally, our results suggest the existence of a regulatory molecule different from the Rsm proteins and dependent on the GacS/GacA (global activator of antibiotic and cyanide production) system, which causes the repression of phaseolotoxin biosynthesis at high temperatures

Evaluación de la capacidad de inhibición de hemólisis oxidativa y actividad antimicrobiana de fracciones peptídicas obtenidas de la hidrólisis de proteínas de huevo, leche y soya usando proteasas extraídas de Bromelia pinguin y Bromelia karatas

Protein hydrolysates are a source of bioactive peptides (BP). They can exert an important role in human health due to their different biological activities. The use of plant proteases is a potential alternative to produce BP. The aim of this work was to evaluate the oxidative hemolytic inhibition capacity (OHI) and antimicrobial activity of hydrolysates and peptide fractions from egg, milk and soy protein hydrolysis with proteases extracted from Bromelia pinguin and Bromelia karatas fruits. In general, all hydrolysates and peptide fractions presented a high OHI, being the peptide fraction of ≤1 kDa from ovalbumin protein hydrolysis with B. karatas proteases that had the greater OHI (98.19%). In contrast, hydrolysates did not register antimicrobial activity, while peptide fractions (≤5, ≤10 y ≤30 kDa), from milk hydrolysis with B. pinguin proteases showed the highest antimicrobial activity against Listeria innocua (22.26-23.79% of inhibition). The results highlight the potential of B. pinguin and B. karatas proteases to hidrolize proteins from different foods, as well as to produce hydrolysates and BP, with high oxidative hemolytic inhibition capacity.Los hidrolizados proteínicos son una fuente de péptidos bioactivos (PB) y estos compuestos pueden ejercer un papel importante en la salud humana debido a sus diferentes acividades biológicas. El uso de proteasas de origen vegetal es una alternativa potencial para producir PB. El objetivo de este trabajo fue evaluar la capacidad de inhibición de la hemólisis oxidativa (IHO) y actividad antimicrobiana de hidrolizados y fracciones peptídicas obtenidas de la hidrólisis de proteínas de huevo (ovoalbúmina), leche y soya, con proteasas extraídas de frutos de Bromelia pinguin y Bromelia karatas. En general, todos los hidrolizados y fracciones peptídicas presentaron una alta IHO, siendo la fracción peptídica de ≤1 kDa, obtenida de la hidrólisis de ovoalbúmina con proteasas de B. karatas, la que presentó mayor actividad (98.19%). En contraste, ninguno de los hidrolizados registró actividad antimicrobiana, mientras que las fracciones peptídicas (≤5, ≤10 y ≤30 kDa), obtenidas de la hidrólisis de leche con proteasas de B. pinguin, mostraron la mayor actividad antimicrobina (22.26-23.79% de inhibición), contra Listeria innocua. Los resultados resaltan el potencial de las proteasas de B. pinguin y B. karatas para hidrolizar proteínas de diferentes alimentos y generar hidrolizados y PB con alta actividad antioxidante

Functional Characterization of the Gene Cluster from Pseudomonas syringae pv. phaseolicola NPS3121 Involved in Synthesis of Phaseolotoxin

Pseudomonas syringae pv. phaseolicola is the causal agent of halo blight disease of beans (Phaseolus vulgaris L.), which is characterized by water-soaked lesions surrounded by a chlorotic halo resulting from the action of a non-host-specific toxin known as phaseolotoxin. This phytotoxin inhibits the enzyme ornithine carbamoyltransferase involved in arginine biosynthesis. Different evidence suggested that genes involved in phaseolotoxin production were clustered. Two genes had been previously identified in our laboratory within this cluster: argK, which is involved in the immunity of the bacterium to its own toxin, and amtA, which is involved in the synthesis of homoarginine. We sequenced the region around argK and amtA in P. syringae pv. phaseolicola NPS3121 to determine the limits of the putative phaseolotoxin gene cluster and to determine the transcriptional pattern of the genes comprising it. We report that the phaseolotoxin cluster (Pht cluster) is composed of 23 genes and is flanked by insertion sequences and transposases. The mutation of 14 of the genes within the cluster lead to a Tox(−) phenotype for 11 of them, while three mutants exhibited low levels of toxin production. The analysis of fusions of selected DNA fragments to uidA, Northern probing, and reverse transcription-PCR indicate the presence of five transcriptional units, two monocistronic and three polycistronic; one is internal to a larger operon. The site for transcription initiation has been determined for each promoter, and the putative promoter regions were identified. Preliminary results also indicate that the gene product of phtL is involved in the regulation of the synthesis of phaseolotoxin