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2018
Pereira, Suzam L. S.; Martins, Cristina P. S.; Sousa, Aurizangela O.; Camillo, Luciana R.; Araújo, Caroline P.; Alcantara, Grazielle M.; Camargo, Danielle S.; Cidade, Luciana C.; Almeida, Alex Alan F. De; Costa, Marcio G. C.
Em: PLOS ONE, vol. 13, iss. 6, pp. e0199187, 2018, ISSN: 1932-6203.
Resumo | Links | BibTeX | Tags: Arabidopsis thaliana, Citrus, Drought adaptation, Gene expression, Genetically modified plants, Leaves, Nicotiana, Oranges
@article{Pereira2018b,
title = {Genome-wide characterization and expression analysis of citrus NUCLEAR FACTOR-Y (NF-Y) transcription factors identified a novel NF-YA gene involved in drought-stress response and tolerance},
author = {Suzam L. S. Pereira and Cristina P. S. Martins and Aurizangela O. Sousa and Luciana R. Camillo and Caroline P. Araújo and Grazielle M. Alcantara and Danielle S. Camargo and Luciana C. Cidade and Alex Alan F. De Almeida and Marcio G. C. Costa},
url = {https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0199187},
doi = {10.1371/JOURNAL.PONE.0199187},
issn = {1932-6203},
year = {2018},
date = {2018-01-01},
journal = {PLOS ONE},
volume = {13},
issue = {6},
pages = {e0199187},
publisher = {Public Library of Science},
abstract = {Nuclear factor Y (NF-Y) is a ubiquitous transcription factor found in eukaryotes. It is composed of three distinct subunits called NF-YA, NF-YB and NF-YC. NF-Ys have been identified as key regulators of multiple pathways in the control of development and tolerance to biotic and abiotic factors. The present study aimed to identify and characterize the complete repertoire of genes coding for NF-Y in citrus, as well as to perform the functional characterization of one of its members, namely CsNFYA5, in transgenic tobacco plants. A total of 22 genes coding for NF-Y were identified in the genomes of sweet orange (Citrus sinensis) and Clementine mandarin (C. clementina), including six CsNF-YAs, 11 CsNF-YBs and five CsNF-YCs. Phylogenetic analyses showed that there is a NF-Y orthologous in the Clementine genome for each sweet orange NF-Y gene; this was not observed when compared to Arabidopsis thaliana. CsNF-Y proteins shared the same conserved domains with their orthologous proteins in other organisms, including mouse. Analysis of gene expression by RNA-seq and EST data demonstrated that CsNF-Ys have a tissue-specific and stress inducible expression profile. qRT-PCR analysis revealed that CsNF-YA5 exhibits differential expression in response to water deficit in leaves and roots of citrus plants. Overexpression of CsNF-YA5 in transgenic tobacco plants contributed to the reduction of H2O2 production under dehydration conditions and increased plant growth and photosynthetic rate under normal conditions and drought stress. These biochemical and physiological responses to drought stress promoted by CsNF-YA5 may confer a productivity advantage in environments with frequent short-term soil water deficit.},
keywords = {Arabidopsis thaliana, Citrus, Drought adaptation, Gene expression, Genetically modified plants, Leaves, Nicotiana, Oranges},
pubstate = {published},
tppubtype = {article}
}
Nuclear factor Y (NF-Y) is a ubiquitous transcription factor found in eukaryotes. It is composed of three distinct subunits called NF-YA, NF-YB and NF-YC. NF-Ys have been identified as key regulators of multiple pathways in the control of development and tolerance to biotic and abiotic factors. The present study aimed to identify and characterize the complete repertoire of genes coding for NF-Y in citrus, as well as to perform the functional characterization of one of its members, namely CsNFYA5, in transgenic tobacco plants. A total of 22 genes coding for NF-Y were identified in the genomes of sweet orange (Citrus sinensis) and Clementine mandarin (C. clementina), including six CsNF-YAs, 11 CsNF-YBs and five CsNF-YCs. Phylogenetic analyses showed that there is a NF-Y orthologous in the Clementine genome for each sweet orange NF-Y gene; this was not observed when compared to Arabidopsis thaliana. CsNF-Y proteins shared the same conserved domains with their orthologous proteins in other organisms, including mouse. Analysis of gene expression by RNA-seq and EST data demonstrated that CsNF-Ys have a tissue-specific and stress inducible expression profile. qRT-PCR analysis revealed that CsNF-YA5 exhibits differential expression in response to water deficit in leaves and roots of citrus plants. Overexpression of CsNF-YA5 in transgenic tobacco plants contributed to the reduction of H2O2 production under dehydration conditions and increased plant growth and photosynthetic rate under normal conditions and drought stress. These biochemical and physiological responses to drought stress promoted by CsNF-YA5 may confer a productivity advantage in environments with frequent short-term soil water deficit.
Kishi, Rosangela Naomi Inui; Stach-Machado, Dagmar; Singulani, Junya Lacorte; Santos, Claudia Tavares; Fusco-Almeida, Ana Marisa; Cilli, Eduardo Maffud; Freitas-Astúa, Juliana; Picchi, Simone Cristina; Machado, Marcos Antonio
Evaluation of cytotoxicity features of antimicrobial peptides with potential to control bacterial diseases of citrus Journal Article
Em: PLOS ONE, vol. 13, iss. 9, pp. e0203451, 2018, ISSN: 1932-6203.
Resumo | Links | BibTeX | Tags: Agrobacterium tumefaciens, Cell membranes, Citrus, Cytotoxicity, Leaves, Lipids, Toxicity, Vesicles
@article{Kishi2018,
title = {Evaluation of cytotoxicity features of antimicrobial peptides with potential to control bacterial diseases of citrus},
author = {Rosangela Naomi Inui Kishi and Dagmar Stach-Machado and Junya Lacorte Singulani and Claudia Tavares Santos and Ana Marisa Fusco-Almeida and Eduardo Maffud Cilli and Juliana Freitas-Astúa and Simone Cristina Picchi and Marcos Antonio Machado},
url = {https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0203451},
doi = {10.1371/JOURNAL.PONE.0203451},
issn = {1932-6203},
year = {2018},
date = {2018-01-01},
journal = {PLOS ONE},
volume = {13},
issue = {9},
pages = {e0203451},
publisher = {Public Library of Science},
abstract = {Antimicrobial peptides (AMPs) can be found in various organisms, and could be considered an alternative for pesticides used to control plant pathogens, including those affecting citrus. Brazil is the largest producer and exporter of frozen concentrated orange juice in the world. However, the citrus industry has been affected by several diseases such as citrus canker and huanglongbing (HLB), caused by the bacteria Xanthomonas citri subsp. citri (X.citri) and Candidatus Liberibacter asiaticus (CaLas), respectively. In order to control these pathogens, putative AMPs were prospected in databases containing citrus sequences. Furthermore, AMPs already reported in the literature were also used for in vitro and in vivo assays against X.citri. Since CaLas cannot be cultivated in vitro, surrogates as Sinorhizobium meliloti and Agrobacterium tumefaciens were used. This study reports the evaluation of six AMPs obtained from different sources, two of them from Citrus spp. (citrus-amp1 and citrus-amp2), three from amphibians (Hylin-a1, K0-W6-Hy-a1 and Ocellatin 4-analogue) and one from porcine (Tritrpticin). Peptides K0-W6-Hy-a1, Ocellatin 4-analogue, and citrus-amp1 showed bactericidal activity against X.citri and S. meliloti and bacteriostatic effect on A. tumefaciens. These results were confirmed for X.citri in planta. In addition cytotoxicity evaluations of these molecules were performed. The AMPs that showed the lowest hemolytic activities were Triptrpticin, citrus-amp1 and citrus-amp2. Citrus-amp1 and citrus-amp2 not presented toxicity in experiments using in vivo model, G. mellonella and U87 MG cells. To verify the interaction of these AMPs with bacteria and erythrocyte cell membranes, vesicles mimicking these cells were built. Citrus-amp1 and Tritrpticin exhibited higher affinity to bacterial membranes, while Ocellatin 4-analogue and Hylin-a1 showed higher affinity to erythrocyte membranes; exclude their use in citrus. This work demonstrates an essential alternative, trough AMPs obtained from Citrus spp., which can be feasibly used to control bacterial pathogens.},
keywords = {Agrobacterium tumefaciens, Cell membranes, Citrus, Cytotoxicity, Leaves, Lipids, Toxicity, Vesicles},
pubstate = {published},
tppubtype = {article}
}
Antimicrobial peptides (AMPs) can be found in various organisms, and could be considered an alternative for pesticides used to control plant pathogens, including those affecting citrus. Brazil is the largest producer and exporter of frozen concentrated orange juice in the world. However, the citrus industry has been affected by several diseases such as citrus canker and huanglongbing (HLB), caused by the bacteria Xanthomonas citri subsp. citri (X.citri) and Candidatus Liberibacter asiaticus (CaLas), respectively. In order to control these pathogens, putative AMPs were prospected in databases containing citrus sequences. Furthermore, AMPs already reported in the literature were also used for in vitro and in vivo assays against X.citri. Since CaLas cannot be cultivated in vitro, surrogates as Sinorhizobium meliloti and Agrobacterium tumefaciens were used. This study reports the evaluation of six AMPs obtained from different sources, two of them from Citrus spp. (citrus-amp1 and citrus-amp2), three from amphibians (Hylin-a1, K0-W6-Hy-a1 and Ocellatin 4-analogue) and one from porcine (Tritrpticin). Peptides K0-W6-Hy-a1, Ocellatin 4-analogue, and citrus-amp1 showed bactericidal activity against X.citri and S. meliloti and bacteriostatic effect on A. tumefaciens. These results were confirmed for X.citri in planta. In addition cytotoxicity evaluations of these molecules were performed. The AMPs that showed the lowest hemolytic activities were Triptrpticin, citrus-amp1 and citrus-amp2. Citrus-amp1 and citrus-amp2 not presented toxicity in experiments using in vivo model, G. mellonella and U87 MG cells. To verify the interaction of these AMPs with bacteria and erythrocyte cell membranes, vesicles mimicking these cells were built. Citrus-amp1 and Tritrpticin exhibited higher affinity to bacterial membranes, while Ocellatin 4-analogue and Hylin-a1 showed higher affinity to erythrocyte membranes; exclude their use in citrus. This work demonstrates an essential alternative, trough AMPs obtained from Citrus spp., which can be feasibly used to control bacterial pathogens.
Pereira, Suzam L. S.; Martins, Cristina P. S.; Sousa, Aurizangela O.; Camillo, Luciana R.; Araújo, Caroline P.; Alcantara, Grazielle M.; Camargo, Danielle S.; Cidade, Luciana C.; Almeida, Alex Alan F. De; Costa, Marcio G. C.
Em: PLOS ONE, vol. 13, iss. 6, pp. e0199187, 2018, ISSN: 1932-6203.
Resumo | Links | BibTeX | Tags: Arabidopsis thaliana, Citrus, Drought adaptation, Gene expression, Genetically modified plants, Leaves, Nicotiana, Oranges
@article{Pereira2018,
title = {Genome-wide characterization and expression analysis of citrus NUCLEAR FACTOR-Y (NF-Y) transcription factors identified a novel NF-YA gene involved in drought-stress response and tolerance},
author = {Suzam L. S. Pereira and Cristina P. S. Martins and Aurizangela O. Sousa and Luciana R. Camillo and Caroline P. Araújo and Grazielle M. Alcantara and Danielle S. Camargo and Luciana C. Cidade and Alex Alan F. De Almeida and Marcio G. C. Costa},
url = {https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0199187},
doi = {10.1371/JOURNAL.PONE.0199187},
issn = {1932-6203},
year = {2018},
date = {2018-01-01},
journal = {PLOS ONE},
volume = {13},
issue = {6},
pages = {e0199187},
publisher = {Public Library of Science},
abstract = {Nuclear factor Y (NF-Y) is a ubiquitous transcription factor found in eukaryotes. It is composed of three distinct subunits called NF-YA, NF-YB and NF-YC. NF-Ys have been identified as key regulators of multiple pathways in the control of development and tolerance to biotic and abiotic factors. The present study aimed to identify and characterize the complete repertoire of genes coding for NF-Y in citrus, as well as to perform the functional characterization of one of its members, namely CsNFYA5, in transgenic tobacco plants. A total of 22 genes coding for NF-Y were identified in the genomes of sweet orange (Citrus sinensis) and Clementine mandarin (C. clementina), including six CsNF-YAs, 11 CsNF-YBs and five CsNF-YCs. Phylogenetic analyses showed that there is a NF-Y orthologous in the Clementine genome for each sweet orange NF-Y gene; this was not observed when compared to Arabidopsis thaliana. CsNF-Y proteins shared the same conserved domains with their orthologous proteins in other organisms, including mouse. Analysis of gene expression by RNA-seq and EST data demonstrated that CsNF-Ys have a tissue-specific and stress inducible expression profile. qRT-PCR analysis revealed that CsNF-YA5 exhibits differential expression in response to water deficit in leaves and roots of citrus plants. Overexpression of CsNF-YA5 in transgenic tobacco plants contributed to the reduction of H2O2 production under dehydration conditions and increased plant growth and photosynthetic rate under normal conditions and drought stress. These biochemical and physiological responses to drought stress promoted by CsNF-YA5 may confer a productivity advantage in environments with frequent short-term soil water deficit.},
keywords = {Arabidopsis thaliana, Citrus, Drought adaptation, Gene expression, Genetically modified plants, Leaves, Nicotiana, Oranges},
pubstate = {published},
tppubtype = {article}
}
Nuclear factor Y (NF-Y) is a ubiquitous transcription factor found in eukaryotes. It is composed of three distinct subunits called NF-YA, NF-YB and NF-YC. NF-Ys have been identified as key regulators of multiple pathways in the control of development and tolerance to biotic and abiotic factors. The present study aimed to identify and characterize the complete repertoire of genes coding for NF-Y in citrus, as well as to perform the functional characterization of one of its members, namely CsNFYA5, in transgenic tobacco plants. A total of 22 genes coding for NF-Y were identified in the genomes of sweet orange (Citrus sinensis) and Clementine mandarin (C. clementina), including six CsNF-YAs, 11 CsNF-YBs and five CsNF-YCs. Phylogenetic analyses showed that there is a NF-Y orthologous in the Clementine genome for each sweet orange NF-Y gene; this was not observed when compared to Arabidopsis thaliana. CsNF-Y proteins shared the same conserved domains with their orthologous proteins in other organisms, including mouse. Analysis of gene expression by RNA-seq and EST data demonstrated that CsNF-Ys have a tissue-specific and stress inducible expression profile. qRT-PCR analysis revealed that CsNF-YA5 exhibits differential expression in response to water deficit in leaves and roots of citrus plants. Overexpression of CsNF-YA5 in transgenic tobacco plants contributed to the reduction of H2O2 production under dehydration conditions and increased plant growth and photosynthetic rate under normal conditions and drought stress. These biochemical and physiological responses to drought stress promoted by CsNF-YA5 may confer a productivity advantage in environments with frequent short-term soil water deficit.
2017
Galdeano, Diogo Manzano; Breton, Michèle Claire; Lopes, João Roberto Spotti; Falk, Bryce W.; Machado, Marcos Antonio
Oral delivery of double-stranded RNAs induces mortality in nymphs and adults of the Asian citrus psyllid, Diaphorina citri Journal Article
Em: PLOS ONE, vol. 12, iss. 3, pp. e0171847, 2017, ISSN: 1932-6203.
Resumo | Links | BibTeX | Tags: Apoptosis, Chitin, Death rates, Diet, Insects, Leaves, Nymphs, RNA interference
@article{Galdeano2017,
title = {Oral delivery of double-stranded RNAs induces mortality in nymphs and adults of the Asian citrus psyllid, Diaphorina citri},
author = {Diogo Manzano Galdeano and Michèle Claire Breton and João Roberto Spotti Lopes and Bryce W. Falk and Marcos Antonio Machado},
url = {https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0171847},
doi = {10.1371/JOURNAL.PONE.0171847},
issn = {1932-6203},
year = {2017},
date = {2017-01-01},
journal = {PLOS ONE},
volume = {12},
issue = {3},
pages = {e0171847},
publisher = {Public Library of Science},
abstract = {The Asian citrus psyllid (ACP), Diaphorina citri Kuwayama, is one of the most important citrus pests. ACP is the vector of the phloem-limited bacteria Candidatus Liberibacter americanus and Candidatus Liberibacter asiaticus, the causal agents of the devastating citrus disease huanglongbing (HLB). The management of HLB is based on the use of healthy young plants, eradication of infected plants and chemical control of the vector. RNA interference (RNAi) has proven to be a promising tool to control pests and explore gene functions. Recently, studies have reported that target mRNA knockdown in many insects can be induced through feeding with double-stranded RNA (dsRNA). In the current study, we targeted the cathepsin D, chitin synthase and inhibitor of apoptosis genes of adult and nymph ACP by feeding artificial diets mixed with dsRNAs and Murraya paniculata leaves placed in dsRNAs solutions, respectively. Adult ACP mortality was positively correlated with the amount of dsRNA used. Both nymphs and adult ACP fed dsRNAs exhibited significantly increased mortality over time compared with that of the controls. Moreover, qRT-PCR analysis confirmed the dsRNA-mediated RNAi effects on target mRNAs. These results showed that RNAi can be a powerful tool for gene function studies in ACP and perhaps for HLB control.},
keywords = {Apoptosis, Chitin, Death rates, Diet, Insects, Leaves, Nymphs, RNA interference},
pubstate = {published},
tppubtype = {article}
}
The Asian citrus psyllid (ACP), Diaphorina citri Kuwayama, is one of the most important citrus pests. ACP is the vector of the phloem-limited bacteria Candidatus Liberibacter americanus and Candidatus Liberibacter asiaticus, the causal agents of the devastating citrus disease huanglongbing (HLB). The management of HLB is based on the use of healthy young plants, eradication of infected plants and chemical control of the vector. RNA interference (RNAi) has proven to be a promising tool to control pests and explore gene functions. Recently, studies have reported that target mRNA knockdown in many insects can be induced through feeding with double-stranded RNA (dsRNA). In the current study, we targeted the cathepsin D, chitin synthase and inhibitor of apoptosis genes of adult and nymph ACP by feeding artificial diets mixed with dsRNAs and Murraya paniculata leaves placed in dsRNAs solutions, respectively. Adult ACP mortality was positively correlated with the amount of dsRNA used. Both nymphs and adult ACP fed dsRNAs exhibited significantly increased mortality over time compared with that of the controls. Moreover, qRT-PCR analysis confirmed the dsRNA-mediated RNAi effects on target mRNAs. These results showed that RNAi can be a powerful tool for gene function studies in ACP and perhaps for HLB control.
Suzuki, Takeshi; España, María Urizarna; Nunes, Maria Andreia; Zhurov, Vladimir; Dermauw, Wannes; Osakabe, Masahiro; Leeuwen, Thomas V. Van; Grbic, Miodrag; Grbic, Vojislava
Protocols for the delivery of small molecules to the two-spotted spider mite, Tetranychus urticae Journal Article
Em: PLOS ONE, vol. 12, iss. 7, pp. e0180658, 2017, ISSN: 1932-6203.
Resumo | Links | BibTeX | Tags: Diet, Eggs, Fluorescent dyes, Larvae, Leaves, Mites, Small molecules, Tracer dye staining
@article{Suzuki2017,
title = {Protocols for the delivery of small molecules to the two-spotted spider mite, Tetranychus urticae},
author = {Takeshi Suzuki and María Urizarna España and Maria Andreia Nunes and Vladimir Zhurov and Wannes Dermauw and Masahiro Osakabe and Thomas V. Van Leeuwen and Miodrag Grbic and Vojislava Grbic},
url = {https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0180658},
doi = {10.1371/JOURNAL.PONE.0180658},
issn = {1932-6203},
year = {2017},
date = {2017-01-01},
journal = {PLOS ONE},
volume = {12},
issue = {7},
pages = {e0180658},
publisher = {Public Library of Science},
abstract = {The two-spotted spider mite, Tetranychus urticae, is a chelicerate herbivore with an extremely wide host range and an extraordinary ability to develop pesticide resistance. Due to its responsiveness to natural and synthetic xenobiotics, the spider mite is becoming a prime pest herbivore model for studies of the evolution of host range, plant-herbivore interactions and mechanisms of xenobiotic resistance. The spider mite genome has been sequenced and its transcriptional responses to developmental and various biotic and abiotic cues have been documented. However, to identify biological and evolutionary roles of T. urticae genes and proteins, it is necessary to develop methods for the efficient manipulation of mite gene function or protein activity. Here, we describe protocols developed for the delivery of small molecules into spider mites. Starting with mite maintenance and the preparation of the experimental mite populations of developmentally synchronized larvae and adults, we describe 3 methods for delivery of small molecules including artificial diet, leaf coating, and soaking. The presented results define critical steps in these methods and demonstrate that they can successfully deliver tracer dyes into mites. Described protocols provide guidelines for high-throughput setups for delivery of experimental compounds that could be used in reverse genetics platforms to modulate gene expression or protein activity, or for screens focused on discovery of new molecules for mite control. In addition, described protocols could be adapted for other Tetranychidae and related species of economic importance such as Varroa, dust and poultry mites.},
keywords = {Diet, Eggs, Fluorescent dyes, Larvae, Leaves, Mites, Small molecules, Tracer dye staining},
pubstate = {published},
tppubtype = {article}
}
The two-spotted spider mite, Tetranychus urticae, is a chelicerate herbivore with an extremely wide host range and an extraordinary ability to develop pesticide resistance. Due to its responsiveness to natural and synthetic xenobiotics, the spider mite is becoming a prime pest herbivore model for studies of the evolution of host range, plant-herbivore interactions and mechanisms of xenobiotic resistance. The spider mite genome has been sequenced and its transcriptional responses to developmental and various biotic and abiotic cues have been documented. However, to identify biological and evolutionary roles of T. urticae genes and proteins, it is necessary to develop methods for the efficient manipulation of mite gene function or protein activity. Here, we describe protocols developed for the delivery of small molecules into spider mites. Starting with mite maintenance and the preparation of the experimental mite populations of developmentally synchronized larvae and adults, we describe 3 methods for delivery of small molecules including artificial diet, leaf coating, and soaking. The presented results define critical steps in these methods and demonstrate that they can successfully deliver tracer dyes into mites. Described protocols provide guidelines for high-throughput setups for delivery of experimental compounds that could be used in reverse genetics platforms to modulate gene expression or protein activity, or for screens focused on discovery of new molecules for mite control. In addition, described protocols could be adapted for other Tetranychidae and related species of economic importance such as Varroa, dust and poultry mites.