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1.
Sci Rep ; 9(1): 8847, 2019 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-31222094

RESUMO

Maternally-transmitted endosymbiotic bacteria are ubiquitous in insects. Among other influential phenotypes, many heritable symbionts of arthropods are notorious for manipulating host reproduction through one of four reproductive syndromes, which are generally exerted during early developmental stages of the host: male feminization; parthenogenesis induction; male killing; and cytoplasmic incompatibility (CI). Major advances have been achieved in understanding mechanisms and identifying symbiont factors involved in reproductive manipulation, particularly male killing and cytoplasmic incompatibility. Nonetheless, whether cytoplasmically-transmitted bacteria influence the maternally-loaded components of the egg or early embryo has not been examined. In the present study, we investigated whether heritable endosymbionts that cause different reproductive phenotypes in Drosophila melanogaster influence the mRNA transcriptome of early embryos. We used mRNA-seq to evaluate differential expression in Drosophila embryos lacking endosymbionts (control) to those harbouring the male-killing Spiroplasma poulsonii strain MSRO-Br, the CI-inducing Wolbachia strain wMel, or Spiroplasma poulsonii strain Hyd1; a strain that lacks a reproductive phenotype and is naturally associated with Drosophila hydei. We found no consistent evidence of influence of symbiont on mRNA composition of early embryos, suggesting that the reproductive manipulation mechanism does not involve alteration of maternally-loaded transcripts. In addition, we capitalized on several available mRNA-seq datasets derived from Spiroplasma-infected Drosophila melanogaster embryos, to search for signals of depurination of rRNA, consistent with the activity of Ribosome Inactivating Proteins (RIPs) encoded by Spiroplasma poulsonii. We found small but statistically significant signals of depurination of Drosophila rRNA in the Spiroplasma treatments (both strains), but not in the symbiont-free control or Wolbachia treatment, consistent with the action of RIPs. The depurination signal was slightly stronger in the treatment with the male-killing strain. This result supports a recent report that RIP-induced damage contributes to male embryo death.


Assuntos
Drosophila melanogaster/embriologia , Drosophila melanogaster/microbiologia , Embrião não Mamífero/microbiologia , Simbiose , Transcriptoma/genética , Animais , Drosophila melanogaster/genética , Feminino , Genes de Insetos/genética , Interações Hospedeiro-Patógeno/genética , Masculino , Fenótipo , RNA Ribossômico , Reprodução/genética , Proteínas Inativadoras de Ribossomos/genética , Proteínas Inativadoras de Ribossomos/fisiologia , Análise de Sequência de RNA , Spiroplasma/enzimologia , Wolbachia
2.
Toxins (Basel) ; 10(4)2018 04 18.
Artigo em Inglês | MEDLINE | ID: mdl-29669991

RESUMO

Ribosome inactivating proteins (RIPs) form a vast family of hundreds of toxins from plants, fungi, algae, and bacteria. RIP activities have also been detected in animal tissues. They exert an N-glycosydase catalytic activity that is targeted to a single adenine of a ribosomal RNA, thereby blocking protein synthesis and leading intoxicated cells to apoptosis. In many cases, they have additional depurinating activities that act against other nucleic acids, such as viral RNA and DNA, or genomic DNA. Although their role remains only partially understood, their functions may be related to plant defense against predators and viruses, plant senescence, or bacterial pathogenesis.


Assuntos
Proteínas Inativadoras de Ribossomos/fisiologia , Toxinas Biológicas/fisiologia , Animais , Humanos , Fenômenos Fisiológicos Vegetais
3.
Molecules ; 20(2): 2364-87, 2015 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-25647575

RESUMO

Sambucus (Adoxaceae) species have been used for both food and medicine purposes. Among these, Sambucus nigra L. (black elder), Sambucus ebulus L. (dwarf elder), and Sambucus sieboldiana L. are the most relevant species studied. Their use has been somewhat restricted due to the presence of bioactive proteins or/and low molecular weight compounds whose ingestion could trigger deleterious effects. Over the last few years, the chemical and pharmacological characteristics of Sambucus species have been investigated. Among the proteins present in Sambucus species both type 1, and type 2 ribosome-inactivating proteins (RIPs), and hololectins have been reported. The biological role played by these proteins remains unknown, although they are conjectured to be involved in defending plants against insect predators and viruses. These proteins might have an important impact on the nutritional characteristics and food safety of elderberries. Type 2 RIPs are able to interact with gut cells of insects and mammals triggering a number of specific and mostly unknown cell signals in the gut mucosa that could significantly affect animal physiology. In this paper, we describe all known RIPs that have been isolated to date from Sambucus species, and comment on their antiviral and entomotoxic effects, as well as their potential uses.


Assuntos
Frutas/química , Extratos Vegetais/farmacologia , Proteínas Inativadoras de Ribossomos/farmacologia , Sambucus/química , Animais , Humanos , Terapia de Alvo Molecular , Extratos Vegetais/isolamento & purificação , Proteínas Inativadoras de Ribossomos/isolamento & purificação , Proteínas Inativadoras de Ribossomos/fisiologia
4.
Plant Sci ; 217-218: 1-7, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24467890

RESUMO

Rice blast caused by Magnaporthe grisea is one of the three major diseases that seriously affect the rice production. Alpha-momorcharin (α-MC), a ribosome-inactivating protein (RIP) isolated from Momordica charantia seeds, has antifungal effects in vitro. In this study, the α-MC gene was constitutively expressed under the control of the 2×35S promoter in transgenic rice (Oryza sativa L.) using an Agrobacterium tumefaciens-mediated method. The nine transgenic plants were obtained and confirmed by PCR and RT-PCR, and the four (B2, B4, B7 and B9) of them whose copy numbers were 1, 2, 3 and 3, respectively, were shown to express the α-MC protein by Western blot. The molecular weight of α-MC in transgenic plants was approximately 38 kDa larger than the purified α-MC protein (28 kDa) in vitro. When the confirmed T1 generations were inoculated with a suspension of M. grisea spores for ten days, the lesions on leaves of transgenic plants were much lesser than those found on wild type (WT). According to the criteria of International Rice Research Institute standard, the mean values for morbidity and disease index numbers were 29.8% and 14.9%, respectively, which were lower than for WT. It is unclear whether RIPs could impact plant fitness and however our results suggest that the α-MC protein is an effective antifungal protein preventing rice blast in transgenic rice.


Assuntos
Interações Hospedeiro-Patógeno , Magnaporthe/fisiologia , Oryza/microbiologia , Proteínas Inativadoras de Ribossomos/fisiologia , Resistência à Doença , Oryza/genética , Oryza/metabolismo , Plantas Geneticamente Modificadas , Proteínas Inativadoras de Ribossomos/genética
5.
Transgenic Res ; 21(4): 785-95, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22038450

RESUMO

Both drought and high salinity stresses are major abiotic factors that limit the yield of agricultural crops. Transgenic techniques have been regarded as effective ways to improve crops in their tolerance to these abiotic stresses. Functional characterization of genes is the prerequisite to identify candidates for such improvement. Here, we have investigated the biological functions of an Oryza sativa Ribosome-inactivating protein gene 18 (OSRIP18) by ectopically expressing this gene under the control of CaMV 35S promoter in the rice genome. We have generated 11 independent transgenic rice plants and all of them showed significantly increased tolerance to drought and high salinity stresses. Global gene expression changes by Microarray analysis showed that more than 100 probe sets were detected with up-regulated expression abundance while signals from only three probe sets were down-regulated after over-expression of OSRIP18. Most of them were not regulated by drought or high salinity stresses. Our data suggested that the increased tolerance to these abiotic stresses in transgenic plants might be due to up-regulation of some stress-dependent/independent genes and OSRIP18 may be potentially useful in further improving plant tolerance to various abiotic stresses by over-expression.


Assuntos
Oryza , Plantas Geneticamente Modificadas , Proteínas Inativadoras de Ribossomos/genética , Estresse Fisiológico/genética , Secas , Regulação da Expressão Gênica de Plantas , Oryza/genética , Oryza/crescimento & desenvolvimento , Oryza/fisiologia , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas/fisiologia , Proteínas Inativadoras de Ribossomos/fisiologia , Tolerância ao Sal/genética
6.
Bone ; 48(6): 1336-45, 2011 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-21356340

RESUMO

Human osteoclast formation from mononuclear phagocyte precursors involves interactions between lectins and their receptors. A type-2 ribosome inactivating protein consists of an A chain and a B chain. The glycosylated B chain binds specifically to galactose moieties of sugar molecules. In this study we showed that the recombinant ribosome inactivating protein B-chain (rRBC) could induce osteoclast formation from human monocytes and murine RAW264.7 macrophages. Tartrate-resistant acid phosphatase (TRAP) staining and bone resorption assays demonstrated that differentiation of osteoclast-like cells was induced in the presence of rRBC in a dose-dependent manner. The rRBC-induced osteoclast differentiation was independent of caspase activation and apoptosis induction activity; however, rRBC-induced osteoclastogenesis was dependent on activation of NF-κB, ERK1/2, and p38 MAP kinase. Thus, our data demonstrated that rRBC induced osteoclast differentiation through a non-apoptotic signaling pathway. In addition to triggering apoptosis, the rRBC also induced osteoclast differentiation. According to this study, a novel role is proposed for rRBC in regulating osteoclast differentiation and in osteoimmunology.


Assuntos
Diferenciação Celular/fisiologia , Macrófagos/citologia , Osteoclastos/citologia , Proteínas Inativadoras de Ribossomos/fisiologia , Animais , Western Blotting , Linhagem Celular , Linhagem da Célula , Ativação Enzimática , Humanos , Sistema de Sinalização das MAP Quinases , Camundongos , Reação em Cadeia da Polimerase Via Transcriptase Reversa
7.
Biochim Biophys Acta ; 1800(2): 190-201, 2010 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-19647040

RESUMO

During the last decade it was unambiguously shown that plants synthesize minute amounts of carbohydrate-binding proteins upon exposure to stress situations like drought, high salt, hormone treatment, pathogen attack or insect herbivory. In contrast to the 'classical' plant lectins, which are typically found in storage vacuoles or in the extracellular compartment this new class of lectins is located in the cytoplasm and the nucleus. Based on these observations the concept was developed that lectin-mediated protein-carbohydrate interactions in the cytoplasm and the nucleus play an important role in the stress physiology of the plant cell. Hitherto, six families of nucleocytoplasmic lectins have been identified. This review gives an overview of our current knowledge on the occurrence of nucleocytoplasmic plant lectins. The carbohydrate-binding properties of these lectins and potential ligands in the nucleocytoplasmic compartment are discussed in view of the physiological role of the lectins in the plant cell.


Assuntos
Estresse Fisiológico/fisiologia , Agaricus/química , Animais , Peptídeos Catiônicos Antimicrobianos/fisiologia , Núcleo Celular/metabolismo , Quimera , Citoplasma/metabolismo , Discoidinas , Proteínas F-Box/fisiologia , Galectinas/fisiologia , Proteínas de Choque Térmico HSP70/fisiologia , Humanos , Lectinas/fisiologia , Ligantes , Lectinas de Ligação a Manose/fisiologia , Lectinas de Plantas/fisiologia , Estrutura Terciária de Proteína , Proteínas de Protozoários/fisiologia , Receptores de Superfície Celular , Proteínas Inativadoras de Ribossomos/fisiologia , Proteínas Inativadoras de Ribossomos Tipo 1
8.
Phytochemistry ; 69(8): 1653-60, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18377939

RESUMO

Pokeweed antiviral protein (PAP) isolated from Phytolacca americana is a ribosome-inactivating protein (RIP) that has RNA N-glycosidase (RNG) activity towards both eukaryotic and prokaryotic ribosomes. In contrast, karasurin-A (KRN), a RIP from Trichosanthes kirilowii var. japonica, is active only on eukaryotic ribosomes. Stepwise selection of chimera proteins between PAP and KRN indicated that the C-terminal region of PAP (residues 209-225) was critical for RNG activity toward prokaryotic ribosomes. When the region of PAP (residues 209-225) was replaced with the corresponding region of KRN the PAP chimera protein, like KRN, was active only on eukaryotic ribosomes. Furthermore, insertion of the region of PAP (residues 209-225) into the KRN chimera protein resulted not only in the detectable RNG activity toward prokaryotic ribosome, but also activity toward the eukaryotic ribosomes as well that was seven-fold higher than for the original KRN. In this study, the possibility of genetic manipulation of the activity and substrate specificity of RIPs is demonstrated.


Assuntos
Células Procarióticas/fisiologia , RNA , Proteínas Inativadoras de Ribossomos Tipo 1/genética , Proteínas Inativadoras de Ribossomos/fisiologia , Ribossomos/fisiologia , Sequência de Aminoácidos , Cristalografia por Raios X , Ativação Enzimática , Modelos Moleculares , Dados de Sequência Molecular , N-Glicosil Hidrolases/química , N-Glicosil Hidrolases/genética , N-Glicosil Hidrolases/fisiologia , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/fisiologia , Conformação Proteica , RNA/química , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Inativadoras de Ribossomos/química , Proteínas Inativadoras de Ribossomos/genética , Proteínas Inativadoras de Ribossomos Tipo 1/química , Proteínas Inativadoras de Ribossomos Tipo 1/fisiologia , Alinhamento de Sequência , Relação Estrutura-Atividade , Especificidade por Substrato
9.
Biochim Biophys Acta ; 1780(5): 773-83, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18222123

RESUMO

Ribosome-inactivating proteins (RIPs, EC 3.2.2.22) are plant enzymes that can inhibit the translation process by removing single adenine residues of the large rRNA. These enzymes are known to function in defense against pathogens, but their biological role is unknown, partly due to the absence of work on RIPs in a model plant. In this study, we purified a protein showing RIP activity from Arabidopsis thaliana by employing chromatography separations coupled with an enzymatic activity. Based on N-terminal and internal amino acid sequencing, the RIP purified was identified as a mature form of pectin methylesterase (PME, At1g11580). The purified native protein showed both PME and RIP activity. PME catalyzes pectin deesterification, releasing acid pectin and methanol, which cause cell wall changes. We expressed the full-length and mature form of cDNA clones into an expression vector and transformed it in Escherichia coli for protein expression. The recombinant PME proteins (full-length and mature) expressed in E. coli did not show either PME or RIP activity, suggesting that post-translational modifications are important for these enzymatic activities. This study demonstrates a new function for an old enzyme identified in a model plant and discusses the possible role of a protein's conformational changes corresponding to its dual enzymatic activity.


Assuntos
Arabidopsis/enzimologia , Hidrolases de Éster Carboxílico/metabolismo , Proteínas Inativadoras de Ribossomos/metabolismo , Sequência de Aminoácidos , Arabidopsis/genética , Sítios de Ligação/genética , Hidrolases de Éster Carboxílico/química , Hidrolases de Éster Carboxílico/genética , Clonagem Molecular , Concentração de Íons de Hidrogênio , Dados de Sequência Molecular , Processamento de Proteína Pós-Traducional , Proteínas Recombinantes/metabolismo , Proteínas Inativadoras de Ribossomos/genética , Proteínas Inativadoras de Ribossomos/fisiologia , Proteínas Inativadoras de Ribossomos Tipo 1/genética , Proteínas Inativadoras de Ribossomos Tipo 1/metabolismo , Proteínas Inativadoras de Ribossomos Tipo 1/fisiologia , Proteínas Inativadoras de Ribossomos Tipo 2/genética , Proteínas Inativadoras de Ribossomos Tipo 2/metabolismo , Proteínas Inativadoras de Ribossomos Tipo 2/fisiologia , Ribossomos/metabolismo , Ricina/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos
10.
Front Biosci ; 13: 3127-35, 2008 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-17981782

RESUMO

Chitinases which catalyze hydrolysis of chitin are believed to be antifungal proteins in plant. Nevertheless, a variety of functions and some new enzymatic activities of chitinases have been found in recent years. We cloned a novel protein from Trichosanthes kirilowii Maximowicz (Family Cucurbitaceae) named TYchi. Expression of TYchi gene in T. kirilowii plants was induced by F. oxysporum, an important cucurbitaceous fungal pathogen, which indicated that TYchi involved in the pathogen-induced plant defense reaction. In addition to its chitin-hydrolytic activity, the recombinant TYchi protein also had RNA N-glycosidase property. In cell-free rabbit reticulocyte lysate system, TYchi inhibited protein synthesis with an IC50 of approximate 5 nM. TYchi also exhibited efficient cytotoxicities to leukemia U937 and choriocarcinoma JAR cells with IC50 about 54 microg ml(-1) and 73 microg ml(-1), respectively. Structure analyses indicated that the putative domain of TYchi is highly similar to the well known active domain of the N-glycosidase trichosanthin (TCS). This bifuntional protein should be useful in diverse applications like RIP-based immunotoxin agent and genetic engineering of plant resistance.


Assuntos
Antineoplásicos/farmacologia , Quitinases/química , Quitinases/fisiologia , Extratos Vegetais/farmacologia , Proteínas de Plantas/farmacologia , Proteínas de Plantas/fisiologia , Proteínas Inativadoras de Ribossomos/fisiologia , Antineoplásicos Fitogênicos , Sistema Livre de Células , Quitinases/antagonistas & inibidores , Clonagem Molecular , Genes de Plantas , Hidrólise , Imageamento Tridimensional , Proteínas de Plantas/química , Plantas , Conformação Proteica , Proteínas Inativadoras de Ribossomos/química , Ribossomos/química , Ribossomos/metabolismo
11.
Artigo em Chinês | MEDLINE | ID: mdl-18349505

RESUMO

Cassin, the new gene of ribosome-inactivating protein (RIP) isolated from Cassia occidentalis, was inserted into expression vector pBI121 to produce plant expression vector pBI121-cassin (Figs.1, 2). pBI121-cassin was introduced into tobacco cultivar 'K326' by the Agrobacteriurm tumefaciens transformation method and more than 100 independent transformants were obtained. Southern blot hybridization analysis showed that a single gene locus was inserted into the chromosome of the transgenic tobacco lines (Fig.5) and PCR analysis of segregation population of progeny indicated that the inheritance of transgene was dominant in transgenic lines (Fig.4, Table 1). Results of RT-PCR and Northern blot hybridization analysis showed that transgene could be transcribed correctly (Figs.5, 6) . Three self-pollination lines of transgenic T(1) and T(2) were challenged with TMV at different concentration titers by mechanical inoculation. The transgenic lines exhibited different levels of resistance to TMV with the nontransgenic plants. After both titers of TMV concentration were inoculated, transgenic lines were considered as the highly resistant type with a delay of 4-13 d in development of symptoms and 10%-25% of test plants were infected, while nontransgenic control plants were susceptible typical symptoms on the newly emerged leaves (Table 2). One T(2) line, T(2)-8-2-1, was regarded as an immune type because it did not show any symptoms during 70 d and all plants were shown to be virus free by ELISA tests.


Assuntos
Nicotiana/genética , Nicotiana/virologia , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/virologia , Proteínas Inativadoras de Ribossomos/fisiologia , Vírus do Mosaico do Tabaco/fisiologia , Agrobacterium tumefaciens/genética , Northern Blotting , Southern Blotting , Cromossomos de Plantas/genética , Proteínas de Plantas/genética , Proteínas de Plantas/fisiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas Inativadoras de Ribossomos/genética
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