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Coronaviruses modify their single-stranded RNA genome with a methylated cap during replication to mimic the eukaryotic mRNAs. The capping process is initiated by several nonstructural proteins (nsp) encoded in the viral genome. The methylation is performed by two methyltransferases, nsp14 and nsp16, while nsp10 acts as a co-factor to both. Additionally, nsp14 carries an exonuclease domain which operates in the proofreading system during RNA replication of the viral genome. Both nsp14 and nsp16 were reported to independently bind nsp10, but the available structural information suggests that the concomitant interaction between these three proteins would be impossible due to steric clashes. Here, we show that nsp14, nsp10, and nsp16 can form a heterotrimer complex upon significant allosteric change. This interaction is expected to encourage the formation of mature capped viral mRNA, modulating nsp14's exonuclease activity, and protecting the viral RNA. Our findings show that nsp14 is amenable to allosteric regulation and may serve as a novel target for therapeutic approaches.
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Metiltransferases , RNA Viral , SARS-CoV-2 , Proteínas não Estruturais Virais , SARS-CoV-2/genética , SARS-CoV-2/metabolismo , Proteínas não Estruturais Virais/metabolismo , Proteínas não Estruturais Virais/genética , Proteínas não Estruturais Virais/química , Metiltransferases/metabolismo , Metiltransferases/genética , Metiltransferases/química , Metilação , RNA Viral/metabolismo , RNA Viral/química , RNA Viral/genética , Exorribonucleases/metabolismo , Exorribonucleases/genética , Humanos , Ligação Proteica , Capuzes de RNA/metabolismo , Capuzes de RNA/genética , Regulação Alostérica , COVID-19/virologia , COVID-19/genética , Multimerização Proteica , Replicação Viral/genética , RNA Mensageiro/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/química , Proteínas Virais Reguladoras e AcessóriasRESUMO
Despite the development of modern drugs, drug resistance in oncology remains the main factor limiting the curability of patients. This paper shows the use of a group of hydrophobic statins to inhibit drug resistance (Pgp protein). In a chemoresistance melanoma cell model, viability, necroptosis with DNA damage, the absorption of the applied pharmaceuticals, and the functional activity of the ABCB1 drug transporter after administration of docetaxel or docetaxel with a selected hydrophobic statin were studied. Taxol-resistant human melanoma cells from three stages of development were used as a model: both A375P and WM239A metastatic lines and radial growth phase WM35 cells. An animal model (Mus musculus SCID) was developed for the A375P cell line. The results show that hydrophobic statins administered with docetaxel increase the accumulation of the drug in the tumor cell a.o. by blocking the ABCB1 channel. They reduce taxol-induced drug resistance. The tumor size reduction was observed after the drug combination was administrated. It was shown that the structural similarity of statins is of secondary importance, e.g., pravastatin and simvastatin. Using cytostatics in the presence of hydrophobic statins increases their effectiveness while reducing their overall toxicity.
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Inibidores de Hidroximetilglutaril-CoA Redutases , Melanoma , Animais , Camundongos , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Docetaxel/farmacologia , Melanoma/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Paclitaxel/farmacologia , Linhagem Celular TumoralRESUMO
Repeated maternal separation (MS) is a useful experimental model in rodents for studying the long-term influence of early-life stress on brain neurophysiology. In our work, we assessed the effect of repeated MS (postnatal day (PND)1-21, 180 min/day) on the postnatal development of rat brain regions involved in memory using proton magnetic resonance spectroscopy (1HMRS) for tissue volume and the level of amino acids such as glutamate, aspartate, glutamine, glycine and gamma-aminobutyric acid (GABA) in the hippocampus. We assessed whether these effects are sex dependent. We also use novel object recognition (NOR) task to examine the effect of MS on memory and the effect of ethanol on it. Finally, we attempted to ameliorate postnatal stress-induced memory deficits by using VU-29, a positive allosteric modulator (PAM) of the metabotropic glutamate type 5 (mGlu5) receptor. In males, we noted deficits in the levels of glutamate, glycine and glutamine and increases in GABA in the hippocampus. In addition, the values of perirhinal cortex, prefrontal cortex and insular cortex and CA3 were decreased in these animals. MS females, in contrast, demonstrated significant increase in glutamate levels and decrease in GABA levels in the hippocampus. Here, the CA1 values alone were increased. VU-29 administration ameliorated these cognitive deficits. Thus, MS stress disturbs amino acids levels mainly in the hippocampus of adult male rats, and enhancement of glutamate neurotransmission reversed recognition memory deficits in these animals.
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Aminoácidos , Disfunção Cognitiva , Feminino , Ratos , Masculino , Animais , Aminoácidos/metabolismo , Glutamina/metabolismo , Caracteres Sexuais , Privação Materna , Encéfalo/metabolismo , Disfunção Cognitiva/metabolismo , Ácido Glutâmico/metabolismo , Ácido gama-Aminobutírico/metabolismo , Transtornos da Memória , Receptor de Glutamato Metabotrópico 5/metabolismo , Hipocampo/metabolismo , Glicina/metabolismoRESUMO
The science related to biomaterials and tissue engineering accounts for a growing part of our knowledge. Surface modifications of biomaterials, their performance in vitro, and the interaction between them and surrounding tissues are gaining more and more attention. It is because we are interested in finding sophisticated materials that help us to treat or mitigate different disorders. Therefore, efficient methods for surface analysis are needed. Several methods are routinely applied to characterize the physical and chemical properties of the biomaterial surface. Mass Spectrometry Imaging (MSI) techniques are able to measure the information about molecular composition simultaneously from biomaterial and adjacent tissue. That is why it can answer the questions connected with biomaterial characteristics and their biological influence. Moreover, this kind of analysis does not demand any antibodies or dyes that may influence the studied items. It means that we can correlate surface chemistry with a biological response without any modification that could distort the image. In our review, we presented examples of biomaterials analyzed by MSI techniques to indicate the utility of SIMS, MALDI, and DESI-three major ones in the field of biomaterials applications. Examples include biomaterials used to treat vascular system diseases, bone implants with the effects of implanted material on adjacent tissues, nanofibers and membranes monitored by mass spectrometry-related techniques, analyses of drug-eluting long-acting parenteral (LAPs) implants and microspheres where MSI serves as a quality control system.
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RATIONALE: Sample preparation is one of the most crucial steps for matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI). Scientists beginning their study with this technique may be overwhelmed by the variety of matrices, solvents, and concentrations; the methods of their applications; and the lack of widely available knowledge of the effect of these parameters on the results. Here we present in depth the aspects of matrix deposition, which will be helpful for the scientific community. METHODS: In this study, we tested several MALDI matrices, such as 2,5-dihydroxybenzoic acid (DHB), norharmane, N-(1-naphthyl)ethylenediamine dihydrochloride (NEDC), and 9-aminoacridine (9AA), using the SunCollect system: wet-interface matrix deposition in the context of lipid analysis. We optimized the number of matrix layers and nozzle settings in terms of spectral intensity and the overall quality of the obtained ion maps. RESULTS: Our research presents the effect of the number of matrix layers and nozzle settings on the results and allows for choosing the optimal parameters for the analyses. In positive ionization mode, DHB matrix could be chosen first. In the negative ionization mode, 1,5-diaminonaphthalene matrix produces a higher peak intensity in a lower mass range and seems to provide more information than 9AA. We recommend NEDC for particular processes such as glucose analysis. Compared to the remaining matrices, norharmane shows significant changes in the obtained ion maps. CONCLUSIONS: Such a large amount of data allow us to observe an interesting conclusion: the obtained ion image for a particular ion could differ dramatically with a change in the matrix, the solvent composition, or even the number of matrix layers. This must be considered when interpreting the result, impelling us to compare the results obtained with different matrices with caution.
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Hidroxibenzoatos , Lipídeos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Solventes , Lipídeos/análise , LasersRESUMO
Imaging mass spectrometry is a powerful technique for the molecular analysis of tissue sections. As in many analytical methods, sample preparation is one of the main and most important steps to obtain results of good quality. Usually, the matrix concentration and solvent composition in different studies are taken for granted without any further consideration. In our studies, we aimed to find how matrix concentration and a type of solvent influence the signal. Moreover, we also aimed to find the relationship between these parameters, how they influence the spectra, and how they influence obtained ion maps. In our experiments, we used SunCollect®, which is a commercially available wet-interface system for matrix deposition. We decided to choose two matrix concentrations (2,5-dihydroxybenzoic acid [DHB]: 15 and 25 mg/mL; 9-aminoacridine [9AA]: 7 and 5 mg/mL) and two different water solutions of solvents in two different percentages for the matrices (DHB: 50% and 70% of methanol [MeOH] and acetonitrile [ACN]; 9AA 70% and 50% of ethanol [EtOH] and MeOH). In the end, the influence of these parameters on obtained spectra and ion maps was assessed.
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Oocytes are a special kind of biological material. Here, the individual variability of a single cell is important. It means that the opportunity to obtain information about the lipid content from the analysis of a single cell is significant. In our study, we present a method for lipid analysis based on the MALDI-based mass spectrometry imaging (MSI) approach. Our attention was paid to the sample preparation optimization with the aid of a wet-interface matrix deposition system (matrix spraying). Technical considerations of the sample preparation process, such as the number of matrix layers and the position of the spraying nozzle during the matrix deposition, are presented in the article. Additionally, we checked if changing the 2,5-dihydroxybenzoic acid (DHB) and 9-Aminoacridine (9AA) matrix concentration and their solvent composition may improve the analysis. Moreover, the comparison of paraformaldehyde-fixed versus nonfixed cell analysis was performed. We hope that our approach will be helpful for those working on lipid analyses in extraordinary material such as a single oocyte. Our study may also offer clues for anybody interested in single-cell analysis with the aid of MALDI mass spectrometry imaging and the wet-interface matrix deposition method.
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The constant increase in the resistance of pathogenic bacteria to the commonly used drugs so far makes it necessary to search for new substances with antibacterial activity. Taking up this challenge, we obtained a series of rhodanine-3-carboxyalkyl acid derivatives containing 2- or 3- or 4-pyridinyl moiety at the C-5 position. These compounds were tested for their antibacterial and antifungal activities. They showed activity against Gram-positive bacteria while they were inactive against Gram-negative bacteria and yeast. In order to explain the relationship between the activity of the compounds and their structure, for selected derivatives crystal structures were determined using the X-ray diffraction method. Modeling of the isosurface of electron density was also performed. For all tested compounds their lipophilicity was determined by the RP-TLC method and by calculation methods. On the basis of the carried-out research, it was found that the derivatives with 1.5 N···S electrostatics interactions between the nitrogen atom in the pyridine moiety and the sulfur atom in the rhodanine system showed the highest biological activity.
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Rodanina , Antibacterianos/química , Antibacterianos/farmacologia , Bactérias Gram-Negativas , Bactérias Gram-Positivas , Testes de Sensibilidade Microbiana , Rodanina/química , Rodanina/farmacologia , Relação Estrutura-AtividadeRESUMO
Semiparametric accelerated failure time (AFT) models are a useful alternative to Cox proportional hazards models, especially when the assumption of constant hazard ratios is untenable. However, rank-based criteria for fitting AFT models are often nondifferentiable, which poses a computational challenge in high-dimensional settings. In this article, we propose a new alternating direction method of multipliers algorithm for fitting semiparametric AFT models by minimizing a penalized rank-based loss function. Our algorithm scales well in both the number of subjects and number of predictors, and can easily accommodate a wide range of popular penalties. To improve the selection of tuning parameters, we propose a new criterion which avoids some common problems in cross-validation with censored responses. Through extensive simulation studies, we show that our algorithm and software is much faster than existing methods (which can only be applied to special cases), and we show that estimators which minimize a penalized rank-based criterion often outperform alternative estimators which minimize penalized weighted least squares criteria. Application to nine cancer datasets further demonstrates that rank-based estimators of semiparametric AFT models are competitive with estimators assuming proportional hazards in high-dimensional settings, whereas weighted least squares estimators are often not. A software package implementing the algorithm, along with a set of auxiliary functions, is available for download at github.com/ajmolstad/penAFT.
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Algoritmos , Modelos Estatísticos , Simulação por Computador , Humanos , Análise dos Mínimos Quadrados , Modelos de Riscos ProporcionaisRESUMO
Recognition of pathogen-associated molecular patterns (PAMPs) by appropriate pattern recognition receptors (PRRs) is a key step in activating the host immune response. The role of a fungal PAMP is attributed to ß-1,3-glucan. The role of α-1,3-glucan, another fungal cell wall polysaccharide, in modulating the host immune response is not clear. This work investigates the potential of α-1,3-glucan as a fungal PAMP by analyzing the humoral immune response of the greater wax moth Galleria mellonella to Aspergillus niger α-1,3-glucan. We demonstrated that 57-kDa and 61-kDa hemolymph proteins, identified as ß-1,3-glucan recognition proteins, bound to A. niger α-1,3-glucan. Other hemolymph proteins, i.e., apolipophorin I, apolipophorin II, prophenoloxidase, phenoloxidase activating factor, arylphorin, and serine protease, were also identified among α-1,3-glucan-interacting proteins. In response to α-1,3-glucan, a 4.5-fold and 3-fold increase in the gene expression of antifungal peptides galiomicin and gallerimycin was demonstrated, respectively. The significant increase in the level of five defense peptides, including galiomicin, corresponded well with the highest antifungal activity in hemolymph. Our results indicate that A. niger α-1,3-glucan is recognized by the insect immune system, and immune response is triggered by this cell wall component. Thus, the role of a fungal PAMP for α-1,3-glucan can be postulated.
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Aspergillus/química , Glucanos/metabolismo , Interações Hospedeiro-Patógeno , Mariposas/microbiologia , Moléculas com Motivos Associados a Patógenos/metabolismo , Animais , Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Corpo Adiposo/efeitos dos fármacos , Corpo Adiposo/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Hemolinfa/metabolismo , Imunização , Larva , Mariposas/efeitos dos fármacos , Mariposas/genética , Ligação Proteica/efeitos dos fármacos , Análise de SobrevidaRESUMO
Mass spectrometry (MS) used in proteomic approaches is able to detect hundreds of proteins in a single assay. Although undeniable high analytical power of MS, data acquired sometimes lead to confusing results, especially during a search of very selective, unique interactions in complex biological matrices. Here, we would like to show an example of such confusing data, providing an extensive discussion on the observed phenomenon. Our investigations focus on the interaction between the Zika virus NS3 protease, which is essential for virus replication. This enzyme is known for helping to remodel the microenvironment of the infected cells. Several reports show that this protease can process cellular substrates and thereby modify cellular pathways that are important for the virus. Herein, we explored some of the targets of NS3, clearly shown by proteomic techniques, as processed during infection. Unfortunately, we could not confirm the biological relevance of protein targets for viral infections detected by MS. Thus, although mass spectrometry is highly sensitive and useful in many instances, also being able to show directions where cell/virus interaction occurs, we believe that deep recognition of their biological role is essential to receive complete insight into the investigated process.
Assuntos
Espectrometria de Massas/métodos , Serina Endopeptidases/metabolismo , Proteínas não Estruturais Virais/metabolismo , Proteínas Virais/metabolismo , Infecção por Zika virus/virologia , Zika virus/metabolismo , Células A549 , Animais , Linhagem Celular , Linhagem Celular Tumoral , Microambiente Celular/fisiologia , Chlorocebus aethiops , Células HEK293 , Humanos , Proteômica/métodos , Transdução de Sinais/fisiologia , Células VeroRESUMO
Mass spectrometry imaging is a powerful tool for analyzing the different kinds of molecules in tissue sections, but some substances cannot be measured easily, due to their physicochemical properties. In such cases, chemical derivatization could be applied to introduce the charge into the molecule and facilitate its detection. Here, we study cholesterol derivatization with betaine aldehyde from tissue slices and evaluate how different sample preparation methods influence the signal from the derivatization product. In this study, we have tested different solutions for betaine aldehyde, different approaches to betaine aldehyde deposition (number of layers, deposition nozzle height), and different MALDI matrices for its analysis. As a result, we proved that the proposed approach could be used for the analysis of cholesterol in different tissues.
Assuntos
Betaína/análogos & derivados , Colesterol/análise , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Animais , Betaína/química , Encéfalo/metabolismo , Cerebelo/metabolismo , Colesterol/química , Íons , Rim/metabolismo , Limite de Detecção , Fígado/metabolismo , CamundongosRESUMO
Type I toxin-antitoxin (TA) systems are widespread genetic modules in bacterial genomes. They express toxic peptides whose overexpression leads to growth arrest or cell death, whereas antitoxins regulate the expression of toxins, acting as labile antisense RNAs. The Staphylococcus aureus (S. aureus) genome contains and expresses several functional type I TA systems, but their biological functions remain unclear. Here, we addressed and challenged experimentally, by proteomics, if the type I TA system, the SprG1/SprF1 pair, influences the overall gene expression in S. aureus. Deleted and complemented S. aureus strains were analyzed for their proteomes, both intracellular and extracellular, during growth. Comparison of intracellular proteomes among the strains points to the SprF1 antitoxin as moderately downregulating protein expression. In the strain naturally expressing the SprG1 toxin, cytoplasmic proteins are excreted into the medium, but this is not due to unspecific cell leakages. Such a toxin-driven release of the cytoplasmic proteins may modulate the host inflammatory response that, in turn, could amplify the S. aureus infection spread.
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Antitoxinas/genética , Proteínas de Bactérias/genética , Regulação Bacteriana da Expressão Gênica/genética , Expressão Gênica/genética , Staphylococcus aureus/genética , Sistemas Toxina-Antitoxina/genética , Citoplasma/genética , Genoma Bacteriano/genética , Proteoma/genética , RNA Antissenso/genéticaRESUMO
Ethanol exposure during pregnancy alters the mammalian target of rapamycin (mTOR) signaling pathway in the fetal brain. Hence, in adult rats exposed to ethanol during the neonatal period, we investigated the influence of rapamycin, an mTOR Complex 1 (mTORC1) inhibitor, on deficits in spatial memory and reversal learning in the Barnes maze task, as well as the ethanol-induced rewarding effects (1.0 or 1.5 g/kg) using the conditioning place preference (CPP) paradigm. Rapamycin (3 and 10 mg/kg) was given before intragastric ethanol (5 g/kg/day) administration at postnatal day (PND)4-9 (an equivalent to the third trimester of human pregnancy). Spatial memory/reversal learning and rewarding ethanol effect were evaluated in adult (PND60-70) rats. Additionally, the impact of rapamycin pre-treatment on the expression of the GluN2B subunit of NMDA receptor in the brain was assessed in adult rats. Our results show that neonatal ethanol exposure induced deficits in spatial memory and reversal learning in adulthood, but the reversal learning outcome may have been due to spatial learning impairments rather than cognitive flexibility impairments. Furthermore, in adulthood the ethanol treated rats were also more sensitive to the rewarding effect of ethanol than the control group. Rapamycin prevented the neonatal effect of ethanol and normalized the GluN2B down-regulation in the hippocampus and the prefrontal cortex, as well as normalized this subunit's up-regulation in the striatum of adult rats. Our results suggest that rapamycin and related drugs may hold promise as a preventive therapy for fetal alcohol spectrum disorders.
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Etanol/toxicidade , Sirolimo/farmacologia , Aprendizagem Espacial/efeitos dos fármacos , Alcoolismo/metabolismo , Animais , Animais Recém-Nascidos/metabolismo , Encéfalo/efeitos dos fármacos , Feminino , Transtornos do Espectro Alcoólico Fetal/tratamento farmacológico , Hipocampo/efeitos dos fármacos , Masculino , Córtex Pré-Frontal/efeitos dos fármacos , Ratos , Ratos Wistar , Receptores de N-Metil-D-Aspartato/genética , Receptores de N-Metil-D-Aspartato/metabolismo , Transdução de Sinais/efeitos dos fármacos , Sirolimo/metabolismo , Aprendizagem Espacial/fisiologiaRESUMO
The work presents identification of antimicrobial peptides and proteins (AMPs) in the hemolymph of Galleria mellonella larvae infected with two Pseudomonas aeruginosa strains (ATCC 27,853 and PA18), differing in the profile of secreted proteases. The insects were immunized with bacteria cultivated in rich (LB) and minimal (M9) media, which resulted in appearance of a similar broad set of AMPs in the hemolymph. Among them, 13 peptides and proteins were identified, i.e. proline-rich peptides 1 and 2, lebocin-like anionic peptide 1 and anionic peptide 2, defensin/galiomicin, cecropin, cecropin D-like peptide, apolipophoricin, gallerimycin, moricin-like peptide B, lysozyme, apolipophorin III, and superoxide dismutase. Bacterial strain- and/or medium-dependent changes in the level of proline-rich peptide 1, anionic peptide 1 and 2, moricin-like peptide B, cecropin D-like and gallerimycin were observed. The analysis of the expression of genes encoding cecropin, gallerimycin, and galiomicin indicated that they were differently affected by the bacterial strain but mainly by the medium used for bacterial culture. The highest expression was found for the LB medium. In addition to the antibacterial and antifungal activity, proteolytic activity was detected in the hemolymph of the P. aeruginosa-infected insects. Based on these results and those presented in our previous reports, it can be postulated that the appearance of AMPs in G. mellonella hemolymph can be triggered not only by P. aeruginosa pathogen associated molecular patterns (PAMPs) but also by bacterial extracellular proteases secreted during infection. However, although there were no qualitative differences in the set of AMPs depending on the P. aeruginosa strain and medium, differences in the level of particular AMPs synthesized in response to the bacteria used were observed.
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Peptídeos Catiônicos Antimicrobianos/metabolismo , Interações Hospedeiro-Patógeno , Mariposas/metabolismo , Peptídeo Hidrolases/metabolismo , Pseudomonas aeruginosa/enzimologia , Animais , Hemolinfa/metabolismo , Larva/metabolismo , Larva/microbiologia , Mariposas/microbiologiaRESUMO
Embryonic diapause (ED) is a temporary arrest of an embryo at the blastocyst stage when it waits for the uterine receptivity signal to implant. ED used by over 100 species may also occur in normally "nondiapausing" mammals when the uterine receptivity signal is blocked or delayed. A large number of lipid droplets (LDs) are stored throughout the preimplantation embryo development, but the amount of lipids varies greatly across different mammalian species. Yet, the role of LDs in the mammalian egg and embryo remains unknown. Here, using a mouse model, we provide evidence that LDs play a crucial role in maintaining ED. By mechanical removal of LDs from zygotes, we demonstrated that delipidated embryos are unable to survive during ED. LDs are not essential for normal prompt implantation, without ED. We further demonstrated that with the progression of ED, the amount of intracellular lipid reduces, and composition changes. This decrease in lipid is caused by a switch from carbohydrate metabolism to lipid catabolism in diapausing blastocysts, which also exhibit increased release of exosomes reflecting elevated embryonic signaling to the mother. We have also shown that presence of LDs in the oocytes of various mammals positively corelates with their species-specific length of diapause. Our results reveal the functional role of LDs in embryonic development. These results can help to develop diagnostic techniques and treatment of recurrent implantation failure and will likely ignite further studies in developmental biology and reproductive medicine fields.
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Blastocisto/metabolismo , Diapausa , Gotículas Lipídicas/metabolismo , Zigoto/metabolismo , Animais , Feminino , CamundongosRESUMO
Red palm weevil (Rhynchophorus ferrugineus Olivier, 1791, Coleoptera: Curculionidae) is a destructive pest of palms, rapidly extending its native geographical range and causing large economic losses worldwide. The present work describes isolation, identification, and bioinformatic analysis of antibacterial proteins and peptides from the immunized hemolymph of this beetle. In total, 17 different bactericidal or bacteriostatic compounds were isolated via a series of high-pressure liquid chromatography steps, and their partial amino acid sequences were determined by N-terminal sequencing or by mass spectrometry. The bioinformatic analysis of the results facilitated identification and description of corresponding nucleotide coding sequences for each peptide and protein, based on the recently published R. ferrugineus transcriptome database. The identified compounds are represented by several well-known bactericidal factors: two peptides similar to defensins, one cecropin-A1-like peptide, and one attacin-B-like protein. Interestingly, we have also identified some unexpected compounds comprising five isoforms of pheromone-binding proteins as well as seven isoforms of odorant-binding proteins. The particular role of these factors in insect response to bacterial infection needs further investigation.
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Antibacterianos/isolamento & purificação , Araceae/parasitologia , Biologia Computacional , Hemolinfa/imunologia , Imunização , Proteínas de Insetos/isolamento & purificação , Peptídeos/isolamento & purificação , Gorgulhos/metabolismo , Sequência de Aminoácidos , Animais , Cromatografia Líquida de Alta Pressão , Cromatografia de Fase Reversa , Proteínas de Insetos/química , Peptídeos/químicaRESUMO
Two series of rhodanine-3-acetic and rhodanine-3-propionic acids derivatives having benzylidene and cinnamylidene substituents with additional electron donating and withdrawing groups at the C-5 position, were synthesised. The structures of the obtained derivatives were confirmed by spectroscopic methods and their lipophilicity was screened. The crystal structures were determined for selected compounds. The antibacterial activity of the derivatives was depended on the type of carboxyalkyl group in the N-3 position and on the type of the substituent in the C-5 position. The derivatives of rhodanine-3-propionic acid demonstrated the highest activity against Gram-positive bacteria. However, none of tested derivatives showed activity against Gram-negative bacteria and yeast. We believe that the presence of the N,N-diethylamine group in the aromatic system and the number of carbon atoms in the carboxyalkyl group is more significant for the biological activity than the fact that the benzylidene or cinnamylidene substituent was present at the C-5 position.
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Reproductive cells are a very special kind of material for the analysis. Depending on the species, their dimensions allow for the application of mass spectrometry imaging-based techniques to receive a reasonable data for interpretation of their condition without any additional sample preparation steps, except for typical sample preparation characteristic for IMS protocols. A comparison between lipid profiles of oocytes could answer the question of the overall quality of the cells in the function of time or conditions of storage. Even tiny differences in the lipid profiles, but still detectable by bioinformatic analysis, could be crucial for the estimation of the conditions of the cells in various stages of development or aging. In our study, MALDI-TOF/TOF MSI was used to analyze and visualize the single oocytes. We deposited the cells on the transparent indium-tin-oxide (ITO) glass and marked their positions, which allowed for the fast localization of the cells and precise laser targeting in the ion source. We also optimized the usage of different MALDI matrices and different approaches. The proposed way of measurement allows analyzing quite a significant quantity of oocytes in a reasonably short time. During the analysis, the lipid composition of the single cell was successfully estimated in a conventional usage of the MALDI ion source, and the localization of lipids was confirmed by imaging mass spectrometry (IMS) analysis. The observed quantity of the lipids allowed for the application of the LIFT™ technique to obtain MS/MS spectra sufficient for lipids' unambiguous identification. We hope that our idea of the oocyte analysis will help to elucidate chemical changes that accompany different processes in which oocytes are involved. There could be such fascinating phenomena as the oocyte maturation, changes in the lipid components during their storage, and much more.
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Virology, as a branch of the life sciences, discovered mass spectrometry (MS) to be the pivotal tool around two decades ago. The technique unveiled the complex network of interactions between the living world of pro- and eukaryotes and viruses, which delivered "a piece of bad news wrapped in protein" as defined by Peter Medawar, Nobel Prize Laureate, in 1960. However, MS is constantly evolving, and novel approaches allow for a better understanding of interactions in this micro- and nanoworld. Currently, we can investigate the interplay between the virus and the cell by analyzing proteomes, interactomes, virus-cell interactions, and search for the compounds that build viral structures. In addition, by using MS, it is possible to look at the cell from the broader perspective and determine the role of viral infection on the scale of the organism, for example, monitoring the crosstalk between infected tissues and the immune system. In such a way, MS became one of the major tools for the modern virology, allowing us to see the infection in the context of the whole cell or the organism. © 2019 John Wiley & Sons Ltd. Mass Spec Rev.