RESUMO
Apurinic/apyrimidinic endonuclease 1/redox factor-1 (APE1/REF-1) is a multifunctional protein acting on cellular signaling pathways, including DNA repair and redox activities. APE1/REF-1 has emerged as a target for cancer therapy, and its role in breast cancer models would reveal new strategies for cancer therapy. APX2009 is a specific APE1/REF-1 redox inhibitor whose anticancer properties have not been described in breast cancer cells. Here, we investigated the effect of the APX2009 treatment in the breast cancer cell lines MDA-MB-231 and MCF-7. Breast cancer cell lines were cultured, and WST1 and colony formation assays were performed to evaluate cell proliferation. Annexin V-FITC/7-AAD and LDH-Glo™ assays were performed to evaluate cell death. The wound healing assay and Matrigel transwell assay were performed after APX2009 treatment to evaluate the cellular migration and invasion processes, respectively. Our findings demonstrated that APX2009 treatment decreased breast cancer cell proliferative, migratory, and invasive properties. Furthermore, it induced apoptosis in both cell lines. Our study is the first to show the effects of APX2009 treatment on apoptosis in a breast cancer cell. Therefore, this study suggested that APX2009 treatment is a promising anticancer molecule for breast cancer.
RESUMO
Abstract ROS (Reactive Oxygen Species) production is a usual plant reaction to environmental stresses such as allelopathy. Plants possess antioxidant enzymes to scavenge cells and resist against the ROS. This study was conducted to evaluate changes in antioxidant enzymes (CAT, GPX, APX) in radish seedlings in response to allelopathic effect of safflower root and shoot residues grown under normal irrigation and drought stress. Safflower allelopathic effect led to an increase in antioxidant enzymes activities. GPX activity increased more than CAT and APX. Radish seedlings exposed to safflower residue grown under drought stress showed more antioxidant enzymes activities. Root residues enhanced the activities of antioxidant enzymes greater than shoot. Seedlings exposed to root residues grown under drought stress had the highest level of antioxidant enzymes activities.
Assuntos
Raízes de Plantas/efeitos adversos , Carthamus/anatomia & histologia , Raphanus/anatomia & histologia , Alelopatia , Antioxidantes/análiseRESUMO
Ascorbate peroxidases (APX) are class I members of the Peroxidase-Catalase superfamily, a large group of evolutionarily related but rather divergent enzymes. Through mining in public databases, unusual subsets of APX homologs were identified, disclosing the existence of two yet uncharacterized families of peroxidases named ascorbate peroxidase-related (APX-R) and ascorbate peroxidase-like (APX-L). As APX, APX-R harbor all catalytic residues required for peroxidatic activity. Nevertheless, proteins of this family do not contain residues known to be critical for ascorbate binding and therefore cannot use it as an electron donor. On the other hand, APX-L proteins not only lack ascorbate-binding residues, but also every other residue known to be essential for peroxidase activity. Through a molecular phylogenetic analysis performed with sequences derived from basal Archaeplastida, the present study discloses the existence of hybrid proteins, which combine features of these three families. The results here presented show that the prevalence of hybrid proteins varies among distinct groups of organisms, accounting for up to 33% of total APX homologs in species of green algae. The analysis of this heterogeneous group of proteins sheds light on the origin of APX-R and APX-L and suggests the occurrence of a process characterized by the progressive deterioration of ascorbate-binding and catalytic sites towards neofunctionalization.
RESUMO
Preharvest applications of methyl jasmonate (MeJA) have been shown to improve post-harvest fruit quality in strawberry fruit. However, the effectiveness of consecutive field applications at different phenological stages on the reinforcement of the antioxidant capacity remains to be analyzed. To determine the best antioxidant response of strawberry (Fragaria × ananassa 'Camarosa') fruit to different numbers and timing of MeJA applications, we performed three differential preharvest treatments (M1, M2, and M3) consisted of successive field applications of 250 µmol L-1 MeJA at flowering (M3), large green (M2 and M3), and ripe fruit stages (M1, M2, and M3). Then, we analyzed their effects on fruit quality parameters [firmness, skin color, soluble solids content/titratable acidity (SSC/TA) ratio, fruit weight at harvest, and weight loss] along with anthocyanin and proanthocyanidin (PA) accumulation; the antioxidant-related enzymatic activity of catalase (CAT), guaiacol peroxidase (POX), and ascorbate peroxidase (APX); the total flavonoid and phenolic contents, antioxidant capacity, and ascorbic acid content (AAC) during post-harvest storage (0, 24, 48, and 72 h). We also evaluated the effect on lignin, total carbon and nitrogen (%C and N), lipid peroxidation, and C and N isotopes signatures on fruits. Remarkably, the results indicated that MeJA treatment increases anthocyanin and PA contents as well as CAT activity in post-harvest storage, depending on the number of preharvest MeJA applications. Also, M3 fruit showed a higher AAC compared to control at 48 and 72 h. Noticeably, the anthocyanin content and CAT activity were more elevated in M3 treatment comparing with control at all post-harvest times. In turn, APX activity was found higher on all MeJA-treated fruit independent of the number of applications. Unlike, MeJA applications did not generate variations on fruit firmness and weight, lignin contents,% C and N, and in lipid peroxidation and water/nitrogen use efficiency according to C and N isotope discrimination. Finally, we concluded that an increasing number of MeJA applications (M3 treatment) improve anthocyanin, PA, AAC, and CAT activity that could play an essential role against reactive oxygen species, which cause stress that affects fruits during post-harvest storage.
RESUMO
The objectives of this study were to determine the serovar of a collection of Actinobacillus pleuropneumoniae strains within the 3-6-8-15 cross-reacting group and to analyze their phenotypic and genetic properties. Based on the serological tests, forty-seven field strains of Actinobacillus pleuropneumoniae isolated from lungs with pleuropneumonia lesions in Japan and Argentina were found to be serovars belonging to the 3-6-8-15 cross-reacting group. By using a capsule loci-based PCR, twenty-nine (96.7%) and one (3.3%) from Japan were identified as serovars 15 and 8, respectively, whereas seventeen (100%) from Argentina were identified as serovar 8. The findings suggested that serovars 8 and 15 were prevalent within the 3-6-8-15 cross-reacting group, in Argentina and Japan, respectively. Phenotypic analyses revealed that the protein patterns observed on SDS-PAGE and the lipopolysaccharide antigen detected by immunoblotting of the reference and field strains of serovars 8 and 15 were similar to each other. Genetic (16S rDNA, apxIIA, apxIIIA, cps, cpx genes, apx and omlA patterns) analyses revealed that the apxIIA and apxIIIA genes of the field strains of serovars 8 and 15 were similar to those of the reference strains of serovars 3, 4, 6, 8 and 15. The results obtained in the present study may be useful for the development of more effective vaccines against disease caused by A. pleuropneumoniae by including the homologous antigens to the most prevalent serovars in specific geographical areas.
Los objetivos del presente estudio fueron determinar el serovar de una colección de cepas de Actinobacillus pleuropneumoniae pertenecientes al grupo 3, 6, 8, 15 de reacciones cruzadas y analizar sus propiedades fenotípicas y genéticas. En base a técnicas serológicas se determinó que cuarenta y siete cepas de A. pleuropneumoniae aisladas a partir de pulmones con lesiones de pleuroneumonía en Japón y Argentina pertenecen al grupo 3, 6, 8, 15. Mediante el uso de PCR basado en locus capsulares, veintinueve (96.7%) y una (3.3%) de los aislados japoneses fueron identificados como serovar 15 y 8 respectivamente, mientras que diecisiete (100%) de los aislados argentinos resultaron pertenecer al serotipo 8. Este hallazgo sugirió que los serovares 8 y 15 fueron los prevalentes dentro del grupo 3, 6, 8, 15 en Japón y Argentina, respectivamente. El análisis fenotípico reveló que los perfiles proteicos determinados por SDS-PAGE, y de antígenos lipopolisacáridos estudiados por inmunoblot, de las cepas de referencia y de campo de los serovares 8 y 15 fueron similares entre sí. El análisis genético (Í6S rDNA, apxIIA, apxIIA, cps, genes cpx, apx y los perfiles omlA) reveló que los genes apxIIA y apxIIIA de las cepas de campo de los serovares 8 y 15 fueron similares a sus homólogos de las cepas de referencia de los serovares 3, 4, 6, 8 y 15. Los resultados obtenidos en el presente estudio pueden ser útiles para el desarrollo de vacunas más efectivas contra la enfermedad causada por A. pleuropneumoniae, al posibilitar incluir antígenos homólogos a los serovares prevalentes en las áreas geográficas de interés.
Assuntos
Animais , Doenças dos Suínos , Infecções por Actinobacillus , Actinobacillus pleuropneumoniae , Argentina , Suínos , Doenças dos Suínos/genética , Infecções por Actinobacillus/genética , Infecções por Actinobacillus/veterinária , Actinobacillus pleuropneumoniae/genética , JapãoRESUMO
The objectives of this study were to determine the serovar of a collection of Actinobacillus pleuropneumoniae strains within the 3-6-8-15 cross-reacting group and to analyze their phenotypic and genetic properties. Based on the serological tests, forty-seven field strains of Actinobacillus pleuropneumoniae isolated from lungs with pleuropneumonia lesions in Japan and Argentina were found to be serovars belonging to the 3-6-8-15 cross-reacting group. By using a capsule loci-based PCR, twenty-nine (96.7%) and one (3.3%) from Japan were identified as serovars 15 and 8, respectively, whereas seventeen (100%) from Argentina were identified as serovar 8. The findings suggested that serovars 8 and 15 were prevalent within the 3-6-8-15 cross-reacting group, in Argentina and Japan, respectively. Phenotypic analyses revealed that the protein patterns observed on SDS-PAGE and the lipopolysaccharide antigen detected by immunoblotting of the reference and field strains of serovars 8 and 15 were similar to each other. Genetic (16S rDNA, apxIIA, apxIIIA, cps, cpx genes, apx and omlA patterns) analyses revealed that the apxIIA and apxIIIA genes of the field strains of serovars 8 and 15 were similar to those of the reference strains of serovars 3, 4, 6, 8 and 15. The results obtained in the present study may be useful for the development of more effective vaccines against disease caused by A. pleuropneumoniae by including the homologous antigens to the most prevalent serovars in specific geographical areas.
Assuntos
Infecções por Actinobacillus , Actinobacillus pleuropneumoniae , Doenças dos Suínos , Infecções por Actinobacillus/genética , Infecções por Actinobacillus/veterinária , Actinobacillus pleuropneumoniae/genética , Animais , Argentina , Japão , Suínos , Doenças dos Suínos/genéticaRESUMO
Apicomplexa is a large phylum of parasitic protists renowned for significant negative health impacts on humans and livestock worldwide. Despite the prevalence and negative impacts of apicomplexans across many animal groups, relatively little attention has been given to apicomplexan parasites of invertebrates, especially marine invertebrates. Previous work has reported an apicomplexan parasite 'X' (APX), a parasite that has been histologically and ultrastructurally identified from the commercially important flat oyster Ostrea chilensis in New Zealand. This apicomplexan may exacerbate host vulnerability to the infectious disease bonamiosis. In this study, we report 18S rRNA sequences amplified from APX-infected O. chilensis tissues. Phylogenetic analyses clearly established that the 18S sequences were of apicomplexan origin; however, their detailed relationship to known apicomplexan groups is less resolved. Two specific probes, designed from the putative APX 18S rRNA sequence, co-localised with APX cells in in situ hybridisations, further supporting our hypothesis that the 18S sequences were from APX. These sequences will facilitate the future development of inexpensive and sensitive molecular diagnostic tests for APX, thereby assisting research focussed on the biology and ecology of this organism and its role in morbidity and mortality of O. chilensis.
Assuntos
Apicomplexa/classificação , Apicomplexa/genética , Ostrea/parasitologia , RNA Ribossômico 18S/genética , Animais , Sequência de Bases , Nova Zelândia , FilogeniaRESUMO
A aplicação de eliciadores em plantas é utilizada em estudos de fisiologia para compreensão dos mecanismos de defesa ao ataque de herbívoros ou infecção por patógenos. Em virtude disso, foi avaliado o efeito do eliciador exógeno quitosana no sistema antioxidante enzimático de jaborandi (Piper mollicomum Kunth). Foram avaliadas as atividades das enzimas ascorbato peroxidase (APX), catalase (CAT) e superóxido dismutase (SOD) e as concentrações de peróxido de hidrogênio (H2O2) e malonaldeído (MDA), ambas análises para verificar a peroxidação lipídica. O delineamento experimental utilizado foi o de blocos casualizados (DBC), constituído de um fatorial (5x2) composto pelos controles sem quitosana (plantas sem pulverização e plantas pulverizadas apenas com o solvente de diluição da quitosana) e concentrações de quitosana (2,5; 5,0 e 10,0 g L-1) em dois estádios de desenvolvimento foliar (em desenvolvimento e completamente expandida). Nas folhas completamente expandidas, o sistema antioxidante foi mais ativo. A CAT teve maior participação no sequestro de radicais livres, induzido pela aplicação da quitosana em ambos os estádios de desenvolvimento foliar. A APX foi induzida somente nas folhas completamente expandidas e na maior concentração de quitosana. O método do MDA foi melhor para evidenciar a diferença nos teores de peróxido de hidrogênio, em função do estresse induzido pela quitosana. De acordo com os resultados obtidos neste ensaio, pode-se sugerir que, nas plantas de jaborandi, as enzimas antioxidativas são requisitadas em resposta ao eliciador em questão, a quitosana, compondo, assim, o mecanismo de defesa dessas plantas.
Elicitors are used in plant physiology studies for comprehension of defense mechanisms against herbivore attach and pathogen infections. For this reason, the exogenous chitosan effect was tested as an elicitor on Jaborandi (Piper mollicomum Kunth). Thus, it was evaluated the elicitor effect in the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) ,ascorbate peroxidase (APX),hydrogen peroxide (H2O2) and malonaldehyde (MDA) concentrations to estimate the lipid peroxidation. The experimental design was in randomized blocks in a factorial 5x2, being 2 controls (plants without pulverization and plants sprayed only with a diluting solvent chitosan) and 3 chitosan concentrations (2.5; 5.0 and 10.0 g L-1) in 2 leaf development stage (in development and fully expanded). At the fully expanded leaves, the antioxidant system presented higher active. The CAT had greater involvement in the kidnapping of free radicals induced by the application of chitosan in both leaf stages of development. The APX was induced only in fully expanded leaves and the highest concentration of chitosan. The MDA method was better to highlight the difference in hydrogen peroxide content as a function of stress induced by chitosan. According to the results of this test, it can be suggested that the plants Jaborandi antioxidant enzymes are required in response to the elicitor in question, chitosan, thus composing the defense mechanism of these plants.
RESUMO
Elicitors are used in plant physiology studies for comprehension of defense mechanisms against herbivore attach and pathogen infections. For this reason, the exogenous chitosan effect was tested as an elicitor on Jaborandi (Piper mollicomum Kunth). Thus, it was evaluated the elicitor effect in the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) ,ascorbate peroxidase (APX),hydrogen peroxide (H2O2) and malonaldehyde (MDA) concentrations to estimate the lipid peroxidation. The experimental design was in randomized blocks in a factorial 5x2, being 2 controls (plants without pulverization and plants sprayed only with a diluting solvent chitosan) and 3 chitosan concentrations (2.5; 5.0 and 10.0 g L-1) in 2 leaf development stage (in development and fully expanded). At the fully expanded leaves, the antioxidant system presented higher active. The CAT had greater involvement in the kidnapping of free radicals induced by the application of chitosan in both leaf stages of development. The APX was induced only in fully expanded leaves and the highest concentration of chitosan. The MDA method was better to highlight the difference in hydrogen peroxide content as a function of stress induced by chitosan. According to the results of this test, it can be suggested that the plants Jaborandi antioxidant enzymes are required in response to the elicitor in question, chitosan, thus composing the defense mechanism of these plants.(AU)
A aplicação de eliciadores em plantas é utilizada em estudos de fisiologia para compreensão dos mecanismos de defesa ao ataque de herbívoros ou infecção por patógenos. Em virtude disso, foi avaliado o efeito do eliciador exógeno quitosana no sistema antioxidante enzimático de jaborandi (Piper mollicomum Kunth). Foram avaliadas as atividades das enzimas ascorbato peroxidase (APX), catalase (CAT) e superóxido dismutase (SOD) e as concentrações de peróxido de hidrogênio (H2O2) e malonaldeído (MDA), ambas análises para verificar a peroxidação lipídica. O delineamento experimental utilizado foi o de blocos casualizados (DBC), constituído de um fatorial (5x2) composto pelos controles sem quitosana (plantas sem pulverização e plantas pulverizadas apenas com o solvente de diluição da quitosana) e concentrações de quitosana (2,5; 5,0 e 10,0 g L-1) em dois estádios de desenvolvimento foliar (em desenvolvimento e completamente expandida). Nas folhas completamente expandidas, o sistema antioxidante foi mais ativo. A CAT teve maior participação no sequestro de radicais livres, induzido pela aplicação da quitosana em ambos os estádios de desenvolvimento foliar. A APX foi induzida somente nas folhas completamente expandidas e na maior concentração de quitosana. O método do MDA foi melhor para evidenciar a diferença nos teores de peróxido de hidrogênio, em função do estresse induzido pela quitosana. De acordo com os resultados obtidos neste ensaio, pode-se sugerir que, nas plantas de jaborandi, as enzimas antioxidativas são requisitadas em resposta ao eliciador em questão, a quitosana, compondo, assim, o mecanismo de defesa dessas plantas.(AU)
Assuntos
Quitosana , Espécies Reativas de Oxigênio , Radicais LivresRESUMO
Elicitors are used in plant physiology studies for comprehension of defense mechanisms against herbivore attach and pathogen infections. For this reason, the exogenous chitosan effect was tested as an elicitor on Jaborandi (Piper mollicomum Kunth). Thus, it was evaluated the elicitor effect in the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) ,ascorbate peroxidase (APX),hydrogen peroxide (H2O2) and malonaldehyde (MDA) concentrations to estimate the lipid peroxidation. The experimental design was in randomized blocks in a factorial 5x2, being 2 controls (plants without pulverization and plants sprayed only with a diluting solvent chitosan) and 3 chitosan concentrations (2.5; 5.0 and 10.0 g L-1) in 2 leaf development stage (in development and fully expanded). At the fully expanded leaves, the antioxidant system presented higher active. The CAT had greater involvement in the kidnapping of free radicals induced by the application of chitosan in both leaf stages of development. The APX was induced only in fully expanded leaves and the highest concentration of chitosan. The MDA method was better to highlight the difference in hydrogen peroxide content as a function of stress induced by chitosan. According to the results of this test, it can be suggested that the plants Jaborandi antioxidant enzymes are required in response to the elicitor in question, chitosan, thus composing the defense mechanism of these plants.
A aplicação de eliciadores em plantas é utilizada em estudos de fisiologia para compreensão dos mecanismos de defesa ao ataque de herbívoros ou infecção por patógenos. Em virtude disso, foi avaliado o efeito do eliciador exógeno quitosana no sistema antioxidante enzimático de jaborandi (Piper mollicomum Kunth). Foram avaliadas as atividades das enzimas ascorbato peroxidase (APX), catalase (CAT) e superóxido dismutase (SOD) e as concentrações de peróxido de hidrogênio (H2O2) e malonaldeído (MDA), ambas análises para verificar a peroxidação lipídica. O delineamento experimental utilizado foi o de blocos casualizados (DBC), constituído de um fatorial (5x2) composto pelos controles sem quitosana (plantas sem pulverização e plantas pulverizadas apenas com o solvente de diluição da quitosana) e concentrações de quitosana (2,5; 5,0 e 10,0 g L-1) em dois estádios de desenvolvimento foliar (em desenvolvimento e completamente expandida). Nas folhas completamente expandidas, o sistema antioxidante foi mais ativo. A CAT teve maior participação no sequestro de radicais livres, induzido pela aplicação da quitosana em ambos os estádios de desenvolvimento foliar. A APX foi induzida somente nas folhas completamente expandidas e na maior concentração de quitosana. O método do MDA foi melhor para evidenciar a diferença nos teores de peróxido de hidrogênio, em função do estresse induzido pela quitosana. De acordo com os resultados obtidos neste ensaio, pode-se sugerir que, nas plantas de jaborandi, as enzimas antioxidativas são requisitadas em resposta ao eliciador em questão, a quitosana, compondo, assim, o mecanismo de defesa dessas plantas.
RESUMO
Elicitors are used in plant physiology studies for comprehension of defense mechanisms against herbivore attach and pathogen infections. For this reason, the exogenous chitosan effect was tested as an elicitor on Jaborandi (Piper mollicomum Kunth). Thus, it was evaluated the elicitor effect in the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) ,ascorbate peroxidase (APX),hydrogen peroxide (H2O2) and malonaldehyde (MDA) concentrations to estimate the lipid peroxidation. The experimental design was in randomized blocks in a factorial 5x2, being 2 controls (plants without pulverization and plants sprayed only with a diluting solvent chitosan) and 3 chitosan concentrations (2.5; 5.0 and 10.0 g L-1) in 2 leaf development stage (in development and fully expanded). At the fully expanded leaves, the antioxidant system presented higher active. The CAT had greater involvement in the kidnapping of free radicals induced by the application of chitosan in both leaf stages of development. The APX was induced only in fully expanded leaves and the highest concentration of chitosan. The MDA method was better to highlight the difference in hydrogen peroxide content as a function of stress induced by chitosan. According to the results of this test, it can be suggested that the plants Jaborandi antioxidant enzymes are required in response to the elicitor in question, chitosan, thus composing the defense mechanism of these plants.
A aplicação de eliciadores em plantas é utilizada em estudos de fisiologia para compreensão dos mecanismos de defesa ao ataque de herbívoros ou infecção por patógenos. Em virtude disso, foi avaliado o efeito do eliciador exógeno quitosana no sistema antioxidante enzimático de jaborandi (Piper mollicomum Kunth). Foram avaliadas as atividades das enzimas ascorbato peroxidase (APX), catalase (CAT) e superóxido dismutase (SOD) e as concentrações de peróxido de hidrogênio (H2O2) e malonaldeído (MDA), ambas análises para verificar a peroxidação lipídica. O delineamento experimental utilizado foi o de blocos casualizados (DBC), constituído de um fatorial (5x2) composto pelos controles sem quitosana (plantas sem pulverização e plantas pulverizadas apenas com o solvente de diluição da quitosana) e concentrações de quitosana (2,5; 5,0 e 10,0 g L-1) em dois estádios de desenvolvimento foliar (em desenvolvimento e completamente expandida). Nas folhas completamente expandidas, o sistema antioxidante foi mais ativo. A CAT teve maior participação no sequestro de radicais livres, induzido pela aplicação da quitosana em ambos os estádios de desenvolvimento foliar. A APX foi induzida somente nas folhas completamente expandidas e na maior concentração de quitosana. O método do MDA foi melhor para evidenciar a diferença nos teores de peróxido de hidrogênio, em função do estresse induzido pela quitosana. De acordo com os resultados obtidos neste ensaio, pode-se sugerir que, nas plantas de jaborandi, as enzimas antioxidativas são requisitadas em resposta ao eliciador em questão, a quitosana, compondo, assim, o mecanismo de defesa dessas plantas.
RESUMO
The main goal of this work was to obtain an orally administered immunogen that would protect against infections by Actinobacillus pleuropneumoniae. The Apx I, II and III toxins were obtained from the supernatants of cultures of serotypes 1 and 3 of A. pleuropneumoniae. The capacity of monoolein gel to trap and protect the Apx toxins, and the effect of their incorporation on the stability of the cubic phase were evaluated. The gel was capable of trapping a 400-µg/ml concentration of the antigen with no effects on its structure. Approximately 60% of the protein molecules were released from the gel within 4h. Four experimental groups were formed, each one with four pigs. All challenges were conducted in a nebulization chamber. Group A: Control (-) not vaccinated and not challenged; Group B: Control (+) not vaccinated but challenged; Group C: vaccinated twice intramuscularly with ToxCom (a commercial toxoid) at an interval of 15 days and then challenged; and Group D: vaccinated orally twice a week for 4 weeks with ToxOral (an oral toxoid) and challenged on day 28 of the experiment with a same dose of 2.0 × 10(4) UFC of A. pleuropneumoniae serotypes 1 and 3. The lesions found in group B covered 27.7-43.1% of the lungs; the pigs in group C had lesions over 12.3-28%; and those in group D over 15.4-32.3%. No lesions were found in the Group A pigs. A. pleuropneumoniae induced macroscopic lesions characteristic of infection by and lesions microscopic detected by histopathology. The etiologic agent was recovered from the infected lungs, tonsils and spleen. The serotypes identified were 1 and 3. An indirect ELISA test identified the antibodies against the Apx toxins in the serum of the animals immunized orally.
Assuntos
Infecções por Actinobacillus/veterinária , Actinobacillus pleuropneumoniae/imunologia , Toxinas Bacterianas/imunologia , Portadores de Fármacos/administração & dosagem , Glicerídeos/administração & dosagem , Pleuropneumonia/veterinária , Doenças dos Suínos/prevenção & controle , Infecções por Actinobacillus/prevenção & controle , Administração Oral , Animais , Toxinas Bacterianas/administração & dosagem , Toxinas Bacterianas/isolamento & purificação , Histocitoquímica , Imunização/métodos , Pulmão/microbiologia , Pulmão/patologia , Tonsila Palatina/microbiologia , Pleuropneumonia/prevenção & controle , Baço/microbiologia , SuínosRESUMO
The inactivation of the chloroplast ascorbate peroxidases (chlAPXs) has been thought to limit the efficiency of the water-water cycle and photo-oxidative protection under stress conditions. In this study, we have generated double knockdown rice (Oryza sativa L.) plants in both OsAPX7 (sAPX) and OsAPX8 (tAPX) genes, which encode chloroplastic APXs (chlAPXs). By employing an integrated approach involving gene expression, proteomics, biochemical and physiological analyses of photosynthesis, we have assessed the role of chlAPXs in the regulation of the protection of the photosystem II (PSII) activity and CO2 assimilation in rice plants exposed to high light (HL) and methyl violagen (MV). The chlAPX knockdown plants were affected more severely than the non-transformed (NT) plants in the activity and structure of PSII and CO2 assimilation in the presence of MV. Although MV induced significant increases in pigment content in the knockdown plants, the increases were apparently not sufficient for protection. Treatment with HL also caused generalized damage in PSII in both types of plants. The knockdown and NT plants exhibited differences in photosynthetic parameters related to efficiency of utilization of light and CO2. The knockdown plants overexpressed other antioxidant enzymes in response to the stresses and increased the GPX activity in the chloroplast-enriched fraction. Our data suggest that a partial deficiency of chlAPX expression modulate the PSII activity and integrity, reflecting the overall photosynthesis when rice plants are subjected to acute oxidative stress. However, under normal growth conditions, the knockdown plants exhibit normal phenotype, biochemical and physiological performance.
Assuntos
Ascorbato Peroxidases/genética , Proteínas de Cloroplastos/genética , Oryza/genética , Estresse Oxidativo/fisiologia , Fotossíntese/genética , Proteínas de Plantas/genética , Ascorbato Peroxidases/metabolismo , Proteínas de Cloroplastos/metabolismo , Eletroforese em Gel Bidimensional , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica/efeitos da radiação , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos da radiação , Herbicidas/farmacologia , Isoenzimas/genética , Isoenzimas/metabolismo , Luz , Oryza/efeitos dos fármacos , Oryza/efeitos da radiação , Estresse Oxidativo/efeitos da radiação , Paraquat/farmacologia , Fotossíntese/efeitos dos fármacos , Fotossíntese/efeitos da radiação , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
The physiological responses of C4 species to simultaneous water deficit and low substrate temperature are poorly understood, as well as the recovery capacity. This study investigated whether the effect of these abiotic stressors is cultivar-dependent. The differential responses of drought-resistant (IACSP94-2094) and drought-sensitive (IACSP97-7065) sugarcane cultivars were characterized to assess the relationship between photosynthesis and antioxidant protection by APX and SOD isoforms under stress conditions. Our results show that drought alone or combined with low root temperature led to excessive energetic pressure at the PSII level. Heat dissipation was increased in both genotypes, but the high antioxidant capacity due to higher SOD and APX activities was genotype-dependent and it operated better in the drought-resistant genotype. High SOD and APX activities were associated with a rapid recovery of photosynthesis in IACSP94-2094 plants after drought and low substrate temperature alone or simultaneously.
Assuntos
Ascorbato Peroxidases/genética , Temperatura Baixa , Secas , Fotossíntese/genética , Saccharum/genética , Superóxido Dismutase/genética , Água , Adaptação Fisiológica/genética , Antioxidantes/metabolismo , Ascorbato Peroxidases/metabolismo , Genótipo , Fenótipo , Complexo de Proteína do Fotossistema II/metabolismo , Transpiração Vegetal , Saccharum/enzimologia , Saccharum/metabolismo , Estresse Fisiológico/genética , Superóxido Dismutase/metabolismoRESUMO
The level of hydrogen peroxide (H2O2) in plants signalizes the induction of several genes, including that of ascorbate peroxidase (APX-EC 1.11.1.11). APX isoenzymes play a central role in the elimination of intracellular H2O2 and contribute to plant responses to diverse stresses. During the infection process in Theobroma cacao by Moniliophthora perniciosa oxidative stress is generated and the APX action recruited from the plant. The present work aimed to characterize the T. cacao APX involved in the molecular interaction of T. cacao-M. perniciosa. The peroxidase activity was analyzed in protein extracts from cocoa plants infected by M. perniciosa and showed the induction of peroxidases like APX in resistant cocoa plants. The cytosolic protein of T. cacao (GenBank: ABR68691.2) was phylogenetically analyzed in relation to other peroxidases from the cocoa genome and eight genes encoding APX proteins with conserved domains were also analyzed. The cDNA from cytosolic APX was cloned in pET28a and the recombinant protein expressed and purified (rTc-cAPX). The secondary structure of the protein was analyzed by Circular Dichroism (CD) displaying high proportion of α-helices when folded. The enzymatic assay shows stable activity using ascorbate and guaiacol as an electron donor for H2O2 reduction. The pH 7.5 is the optimum for enzyme activity. Chromatographic analysis suggests that rTc-cAPX is a homodimer in solution. Results indicate that the rTc-cAPX is correctly folded, stable and biochemically active. The purified rTc-cAPX presented biotechnological potential and is adequate for future structural and functional studies.
Assuntos
Agaricales , Ascorbato Peroxidases , Cacau , Resistência à Doença , Estresse Oxidativo , Doenças das Plantas/microbiologia , Proteínas de Plantas , Ascorbato Peroxidases/química , Ascorbato Peroxidases/genética , Ascorbato Peroxidases/metabolismo , Cacau/enzimologia , Cacau/genética , Cacau/microbiologia , Citosol , DNA Complementar , Dimerização , Resistência à Doença/genética , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Peróxido de Hidrogênio/metabolismo , Concentração de Íons de Hidrogênio , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Dobramento de Proteína , Estrutura Secundária de Proteína , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismoRESUMO
Arsenate (As(V)) and arsenite (As(III)) contamination is able to interfere negatively on plant metabolism, promoting a reduction of nutrients uptake and transport and also an increase of reactive oxygen species (ROS) generation. However, some plants are considered tolerant against As exposure through the activation of defense mechanisms. Therefore, this study aimed to evaluate the effects of different As(V) and As(III) concentrations (0.0, 6.6, 13.2, 26.4 and 52.8 µmol L(-1)), on mineral nutrients concentration [calcium (Ca), magnesium (Mg), phosphorous (P), iron (Fe), manganese (Mg) and copper (Cu)], on membrane lipid peroxidation and also on the enzymes belonging to the antioxidant defense system [superoxide dismutase (SOD), total peroxidase (POX), catalase (CAT), glutathione reductase (GR) and ascorbate peroxidase (APX)] of plants of Lactuca sativa L. cv Hanson. As(V) and As(III), showed, in general, the same toxic effects in leaves and roots with significant changes in essential macro- and micronutrients concentration. Lipid peroxidation of cellular membranes was also observed in tested plants, probably resulted from an action of ROS generated by this metalloid. The increase of ROS generation and their scavenge were evident since an increase of SOD, POX, CAT and APX activity in leaves, and SOD, CAT and GR activity in roots were observed. Therefore, As(V) and As(III) exposure resulted in toxic effects in leaves and roots of lettuce plants; however, this plant species was able to attenuate these potential As damages through the activation of defense mechanisms, keeping its metabolism. Arsenic-tolerant plants are considered a great risk to the public health since it results in As insertion to the food chain.
Assuntos
Arseniatos/toxicidade , Arsenitos/toxicidade , Lactuca/efeitos dos fármacos , Lactuca/metabolismo , Antioxidantes/metabolismo , Ascorbato Peroxidases/metabolismo , Catalase/metabolismo , Glutationa Redutase/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Peroxidase/metabolismo , Superóxido Dismutase/metabolismoRESUMO
Identification of metabolic targets of environmental stress factors is critical to improve the stress tolerance of plants. Studying the biochemical and physiological responses of plants with different capacities to deal with stress is a valid approach to reach this objective. Lotus corniculatus (lotus) and Trifolium pratense (clover) are legumes with contrasting summer stress tolerances. In stress conditions, which are defined as drought, heat or a combination of both, we found that differential biochemical responses of leaves explain these behaviours. Lotus and clover showed differences in water loss control, proline accumulation and antioxidant enzymatic capacity. Drought and/or heat stress induced a large accumulation of proline in the tolerant species (lotus), whereas heat stress did not cause proline accumulation in the sensitive species (clover). In lotus, Mn-SOD and Fe-SOD were induced by drought, but in clover, the SOD-isoform profile was not affected by stress. Moreover, lotus has more SOD-isoforms and a higher total SOD activity than clover. The functionality and electrophoretic profile of photosystem II (PSII) proteins under stress also exhibited differences between the two species. In lotus, PSII activity was drastically affected by combined stress and, interestingly, was correlated with D2 protein degradation. Possible implications of this event as an adaption mechanism in tolerant species are discussed. We conclude that the stress-tolerant capability of lotus is related to its ability to respond to oxidative damage and adaption of the photosynthetic machinery. This reveals that these two aspects should be included in the evaluation of the tolerance of species to stress conditions.
Assuntos
Adaptação Fisiológica , Secas , Temperatura Alta , Lotus/fisiologia , Oxirredutases/metabolismo , Complexo de Proteína do Fotossistema II/metabolismo , Trifolium/fisiologia , Antioxidantes , Lotus/enzimologia , Lotus/metabolismo , Proteínas de Plantas/metabolismo , Prolina/metabolismo , Estresse Fisiológico , Superóxido Dismutase/metabolismo , Trifolium/enzimologia , Trifolium/metabolismo , ÁguaRESUMO
Tryparedoxins (TXNs) are multipurpose oxidoreductases from trypanosomatids that transfer reducing equivalents from trypanothione to various thiol proteins. In Trypanosoma cruzi, two genes coding for TXN-like proteins have been identified: TXNI, previously characterized as a cytoplasmic protein, and TXNII, a putative tail-anchored membrane protein. In this work, we performed a comparative functional characterization of T. cruzi TXNs. Particularly, we cloned the gene region coding for the soluble version of TXNII for its heterologous expression. The truncated recombinant protein (without its 22 C-terminal transmembrane amino acids) showed TXN activity. It was also able to transfer reducing equivalents from trypanothione, glutathione, or dihydrolipoamide to various acceptors, including methionine sulfoxide reductases and peroxiredoxins. The results support the occurrence and functionality of a second tryparedoxin, which appears as a new component in the redox scenario for T. cruzi.