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1.
Chem Commun (Camb) ; 57(72): 9096-9099, 2021 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-34498651

RESUMO

We present a detailed computational analysis of the binding mode and reactivity of the novel oral inhibitor PF-07321332 developed against the SARS-CoV-2 3CL protease. Alchemical free energy calculations suggest that positions P3 and P4 could be susceptible to improvement in order to get a larger binding strength. QM/MM simulations unveil the reaction mechanism for covalent inhibition, showing that the nitrile warhead facilitates the recruitment of a water molecule for the proton transfer step.


Assuntos
Proteases 3C de Coronavírus/antagonistas & inibidores , Simulação de Dinâmica Molecular , Nitrilas/química , Inibidores de Proteases/química , SARS-CoV-2/enzimologia , Sítios de Ligação , COVID-19/patologia , COVID-19/virologia , Domínio Catalítico , Proteases 3C de Coronavírus/metabolismo , Humanos , Lactamas/química , Lactamas/metabolismo , Leucina/química , Leucina/metabolismo , Nitrilas/metabolismo , Prolina/química , Prolina/metabolismo , Inibidores de Proteases/metabolismo , Teoria Quântica , SARS-CoV-2/isolamento & purificação , Termodinâmica
2.
Int J Mol Sci ; 22(15)2021 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-34360688

RESUMO

Plant response to osmotic stress is a complex issue and includes a wide range of physiological and biochemical processes. Extensive studies of known cultivars and their reaction to drought or salinity stress are very important for future breeding of new and tolerant cultivars. Our study focused on the antioxidant activity, accumulations of osmotica, and the content of abscisic acid in apple (cv. "Malinové holovouské", "Fragrance", "Rubinstep", "Idared", "Car Alexander") and cherry (cv. "Regina", "Napoleonova", "Kastánka", "Sunburst", "P-HL-C") cultivated in vitro on media containing different levels of polyethylene glycol PEG-6000. Our results indicated that the studied genotypes responded differently to osmotic stress manifested as reduction in the leaf relative water content (RWC) and increment in the activities of antioxidant enzymes, proline, sugars, and abscisic acid content. Overall, cherry cultivars showed a smaller decrease in percentage RWC and enzymatic activities, but enhanced proline content compared to the apple plants cultivars. Cultivars "Rubinstep", "Napoleonova", and "Kastánka" exhibited higher antioxidant capacity and accumulation of osmoprotectants like proline and sorbitol that can be associated with the drought-tolerance system.


Assuntos
Ácido Abscísico/análise , Antioxidantes/análise , Pressão Osmótica , Prolina/análise , Estresse Fisiológico , Açúcares/análise , Malus/química , Malus/metabolismo , Malus/fisiologia , Prolina/metabolismo , Prunus avium/química , Prunus avium/metabolismo , Prunus avium/fisiologia , Açúcares/metabolismo
3.
BMC Plant Biol ; 21(1): 368, 2021 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-34384391

RESUMO

BACKGROUND: Melatonin is considered a potential plant growth regulator to enhance the growth of plants and increase tolerance to various abiotic stresses. Nevertheless, melatonin's role in mediating stress response in different plant species and growth cycles still needs to be explored. This study was conducted to understand the impact of different melatonin concentrations (0, 50, 100, and 150 µM) applied as a soil drench to maize seedling under drought stress conditions. A decreased irrigation approach based on watering was exposed to maize seedling after drought stress was applied at 40-45% of field capacity. RESULTS: The results showed that drought stress negatively affected the growth behavior of maize seedlings, such as reduced biomass accumulation, decreased photosynthetic pigments, and enhanced the malondialdehyde and reactive oxygen species (ROS). However, melatonin application enhanced plant growth; alleviated ROS-induced oxidative damages by increasing the photosynthetic pigments, antioxidant enzyme activities, relative water content, and osmo-protectants of maize seedlings. CONCLUSIONS: Melatonin treatment also enhanced the stomatal traits, such as stomatal length, width, area, and the number of pores under drought stress conditions. Our data suggested that 100 µM melatonin application as soil drenching could provide a valuable foundation for improving plant tolerance to drought stress conditions.


Assuntos
Melatonina/farmacologia , Reguladores de Crescimento de Plantas/farmacologia , Zea mays/efeitos dos fármacos , Zea mays/crescimento & desenvolvimento , Antioxidantes/metabolismo , Biomassa , Clorofila/metabolismo , Secas , Estresse Oxidativo , Fotossíntese/efeitos dos fármacos , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/ultraestrutura , Proteínas de Plantas/metabolismo , Estômatos de Plantas/efeitos dos fármacos , Estômatos de Plantas/ultraestrutura , Prolina/metabolismo , Espécies Reativas de Oxigênio , Plântula/efeitos dos fármacos , Plântula/crescimento & desenvolvimento , Açúcares/metabolismo , Zea mays/enzimologia , Zea mays/fisiologia
4.
Int J Mol Sci ; 22(16)2021 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-34445791

RESUMO

Collagen is heavily hydroxylated. Experiments show that proline hydroxylation is important to triple helix (monomer) stability, fibril assembly, and interaction of fibrils with other molecules. Nevertheless, experiments also show that even without hydroxylation, type I collagen does assemble into its native D-banded fibrillar structure. This raises two questions. Firstly, even though hydroxylation removal marginally affects macroscopic structure, how does such an extensive chemical change, which is expected to substantially reduce hydrogen bonding capacity, affect local structure? Secondly, how does such a chemical perturbation, which is expected to substantially decrease electrostatic attraction between monomers, affect collagen's mechanical properties? To address these issues, we conduct a benchmarked molecular dynamics study of rat type I fibrils in the presence and absence of hydroxylation. Our simulations reproduce the experimental observation that hydroxylation removal has a minimal effect on collagen's D-band length. We also find that the gap-overlap ratio, monomer width and monomer length are minimally affected. Surprisingly, we find that de-hydroxylation also has a minor effect on the fibril's Young's modulus, and elastic stress build up is also accompanied by tightening of triple-helix windings. In terms of local structure, de-hydroxylation does result in a substantial drop (23%) in inter-monomer hydrogen bonding. However, at the same time, the local structures and inter-monomer hydrogen bonding networks of non-hydroxylated amino acids are also affected. It seems that it is this intrinsic plasticity in inter-monomer interactions that preclude fibrils from undergoing any large changes in macroscopic properties. Nevertheless, changes in local structure can be expected to directly impact collagen's interaction with extra-cellular matrix proteins. In general, this study highlights a key challenge in tissue engineering and medicine related to mapping collagen chemistry to macroscopic properties but suggests a path forward to address it using molecular dynamics simulations.


Assuntos
Colágeno Tipo I/metabolismo , Hidroxilação/fisiologia , Animais , Módulo de Elasticidade/fisiologia , Ligação de Hidrogênio , Simulação de Dinâmica Molecular , Prolina/metabolismo , Ratos
5.
Nat Commun ; 12(1): 3396, 2021 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-34099711

RESUMO

Amyotrophic lateral sclerosis and frontotemporal dementia are two neurodegenerative diseases with overlapping clinical features and the pathological hallmark of cytoplasmic deposits of misfolded proteins. The most frequent cause of familial forms of these diseases is a hexanucleotide repeat expansion in the non-coding region of the C9ORF72 gene that is translated into dipeptide repeat polymers. Here we show that proline/arginine repeat polymers derail protein folding by sequestering molecular chaperones. We demonstrate that proline/arginine repeat polymers inhibit the folding catalyst activity of PPIA, an abundant molecular chaperone and prolyl isomerase in the brain that is altered in amyotrophic lateral sclerosis. NMR spectroscopy reveals that proline/arginine repeat polymers bind to the active site of PPIA. X-ray crystallography determines the atomic structure of a proline/arginine repeat polymer in complex with the prolyl isomerase and defines the molecular basis for the specificity of disease-associated proline/arginine polymer interactions. The combined data establish a toxic mechanism that is specific for proline/arginine dipeptide repeat polymers and leads to derailed protein homeostasis in C9orf72-associated neurodegenerative diseases.


Assuntos
Esclerose Amiotrófica Lateral/patologia , Proteína C9orf72/genética , Dipeptídeos/metabolismo , Demência Frontotemporal/patologia , Peptidilprolil Isomerase/metabolismo , Esclerose Amiotrófica Lateral/genética , Arginina/genética , Arginina/metabolismo , Biopolímeros/metabolismo , Encéfalo/patologia , Domínio Catalítico , Cristalografia por Raios X , Expansão das Repetições de DNA , Dipeptídeos/genética , Demência Frontotemporal/genética , Humanos , Ressonância Magnética Nuclear Biomolecular , Peptidilprolil Isomerase/isolamento & purificação , Peptidilprolil Isomerase/ultraestrutura , Prolina/genética , Prolina/metabolismo , Agregados Proteicos/genética , Ligação Proteica , Dobramento de Proteína , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/ultraestrutura , Sequências Repetitivas de Aminoácidos/genética
6.
Ecotoxicol Environ Saf ; 220: 112402, 2021 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-34090105

RESUMO

High levels of soil salinity can cause substantial decline in growth and productivity of crops worldwide, thus representing a major threat to global agriculture. In recent years, engineered nanoparticles (NPs) have been deemed as a promising alternative in combating abiotic stress factors, such as salinity. In this context, the present study was designed to explore the potential of cerium oxide nanoparticles (CeO2NPs) in alleviating salt stress in grapevine (Vitis vinifera L. cv. Flame Seedless) cuttings. Specifically, the interaction between CeO2 NPs (25, 50 and 100 mg L-1) and salinity (25 and 75 mM NaCl) was evaluated by assaying an array of agronomic, physiological, analytical and biochemical parameters. Treatments with CeO2 NPs, in general, alleviated the adverse impacts of salt stress (75 mM NaCl) significantly improving relevant agronomic traits of grapevine. CeO2 NPs significantly ameliorated chlorophyll damage under high levels of salinity. Furthermore, the presence of CeO2 NPs attenuated salinity-induced damages in grapevine as indicated by lower levels of proline, MDA and EL; however, H2O2 content was not ameliorated by the presence of CeO2 NPs under salt stress. Additionally, salinity caused substantial increases in enzymatic activities of GP, APX and SOD, compared with control plants. Similar to stress conditions, all concentrations of CeO2 NPs triggered APX activity, while the highest concentration of CeO2 NPs significantly increased GP activity. However, CeO2 NPs did not significantly modify SOD activity. Considering mineral nutrient profile, salinity increased Na and Cl content as well as Na/K ratio, while it decreased K, P and Ca contents. Nevertheless, the presence of CeO2 NPs did not lead to significant alterations in Na, K and P content of salt-stressed plants. Taken together, current findings suggest that CeO2 NPs could be employed as promising salt-stress alleviating agents in grapevine.


Assuntos
Cério/farmacologia , Nanopartículas , Estresse Salino/efeitos dos fármacos , Vitis/efeitos dos fármacos , Antioxidantes/metabolismo , Cério/química , Clorofila/metabolismo , Peróxido de Hidrogênio/metabolismo , Nanopartículas/química , Prolina/metabolismo , Salinidade , Solo/química , Vitis/metabolismo
7.
Int J Mol Sci ; 22(10)2021 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-34065025

RESUMO

A new family of hybrid ß,γ-peptidomimetics consisting of a repetitive unit formed by a chiral cyclobutane-containing trans-ß-amino acid plus a Nα-functionalized trans-γ-amino-l-proline joined in alternation were synthesized and evaluated as cell penetrating peptides (CPP). They lack toxicity on the human tumoral cell line HeLa, with an almost negligible cell uptake. The dodecapeptide showed a substantial microbicidal activity on Leishmania parasites at 50 µM but with a modest intracellular accumulation. Their previously published γ,γ-homologues, with a cyclobutane γ-amino acid, showed a well-defined secondary structure with an average inter-guanidinium distance of 8-10 Å, a higher leishmanicidal activity as well as a significant intracellular accumulation. The presence of a very rigid cyclobutane ß-amino acid in the peptide backbone precludes the acquisition of a defined conformation suitable for their cell uptake ability. Our results unveiled the preorganized charge-display as a relevant parameter, additional to the separation among the charged groups as previously described. The data herein reinforce the relevance of these descriptors in the design of CPPs with improved properties.


Assuntos
Peptídeos Penetradores de Células/metabolismo , Ciclobutanos/metabolismo , Leishmania/metabolismo , Peptidomiméticos/metabolismo , Prolina/metabolismo , Sobrevivência Celular , Peptídeos Penetradores de Células/química , Ciclobutanos/química , Dimerização , Células HeLa , Humanos , Peptidomiméticos/química , Prolina/química , Conformação Proteica
8.
Plant Cell Rep ; 40(7): 1155-1170, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33950277

RESUMO

KEY MESSAGE: The drought and salt tolerances of wheat were enhanced by ectopic expression of the Arabidopsis ornithine aminotransferase (AtOAT) encoded gene. The OAT was confirmed to play a role in proline biosynthesis in wheat. Proline (Pro) accumulation is a common response to both abiotic and biotic stresses in plants. Ornithine aminotransferase (OAT) is pyridoxal-5-phosphate dependent enzyme involved in plant proline biosynthesis. During stress condition, proline is synthesized via glutamate and ornithine pathways. The OAT is the key enzyme in ornithine pathway. In this study, an OAT gene AtOAT from Arabidopsis was expressed in wheat for its functional characterization under drought, salinity, and heat stress conditions. We found that the expression of AtOAT enhanced the drought and salt stress tolerances of wheat by increasing the proline content and peroxidase activity. In addition, it was confirmed that the expression of AtOAT also played a partial tolerance to heat stress in the transgenic wheat plants. Moreover, quantitative real-time PCR (qRT-PCR) analysis showed that the transformation of AtOAT up-regulated the expression of the proline biosynthesis associated genes TaOAT, TaP5CS, and TaP5CR, and down-regulated that of the proline catabolism related gene TaP5CDH in the transgenic plants under stress conditions. Moreover, the genes involved in ornithine pathway (Orn-OAT-P5C/GSA-P5CR-Pro) were up-regulated along with the up-regulation of those genes involved in glutamate pathway (Glu-P5CS-P5C/GSA-P5CR-Pro). Therefore, we concluded that the expression of AtOAT enhanced wheat abiotic tolerance via modifying the proline biosynthesis by up-regulating the expression of the proline biosynthesis-associated genes and down-regulating that of the proline catabolic gene under stresses condition.


Assuntos
Proteínas de Arabidopsis/genética , Ornitina-Oxo-Ácido Transaminase/genética , Plantas Geneticamente Modificadas/fisiologia , Estresse Fisiológico/genética , Triticum/fisiologia , Secas , Regulação da Expressão Gênica de Plantas , Resposta ao Choque Térmico/genética , Plantas Geneticamente Modificadas/genética , Prolina/genética , Prolina/metabolismo , Tolerância ao Sal/genética , Estresse Fisiológico/fisiologia , Triticum/genética
9.
BMC Plant Biol ; 21(1): 233, 2021 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-34034675

RESUMO

BACKGROUND: Lanthanum (La) is a rare earth element that can influence plant growth and development. However, the effect of La on growth, yield formation and 2-acetyl-1-pyrroline (2-AP, a key compound responsible for the aroma of rice) biosynthesis in aromatic rice (Oryza sativa L. subsp. japonica Kato) has not been reported. Therefore, the present study investigated the effects of La on growth, photosynthesis, yield formation and 2-AP biosynthesis in aromatic rice through three experiments. RESULTS: Two pot experiments and a two-year field trial were conducted with different rates of La application (20-120 LaCl3 mg kg-1 and 12 kg ha-1 LaCl3), and treatments without La application were used as controls. The results showed that the application of LaCl3 at 80 and 100 mg kg-1 and at 12 kg ha-1 greatly increased the 2-AP content (by 6.45-43.03%) in aromatic rice seedlings and mature grains compared with the control. The La treatments also increased the chlorophyll content, net photosynthetic rate and total aboveground biomass of rice seedlings. Higher antioxidant enzyme (superoxide, peroxidase, and catalase) activity was detected in the La treatments than in the control. The La treatments also increased the grain yield, grain number per panicle and seed-setting rate of aromatic rice relative to the control. Moreover, the grain proline and γ-aminobutyric acid contents and the activity of betaine aldehyde dehydrogenase significantly decreased under the La treatment. The application of La to soil enhanced the activity of proline dehydrogenase by 20.62-56.95%. CONCLUSIONS: La improved the growth, yield formation and 2-AP content of aromatic rice and enhanced 2-AP biosynthesis by increasing the conversion of proline to 2-AP and decreasing the conversion of GABald to GABA.


Assuntos
Antioxidantes/metabolismo , Lantânio/farmacologia , Oryza/fisiologia , Proteínas de Plantas/metabolismo , Pirróis/metabolismo , Clorofila/metabolismo , Grão Comestível , Oryza/genética , Oryza/crescimento & desenvolvimento , Fotossíntese , Proteínas de Plantas/genética , Prolina/metabolismo , Prolina Oxidase/genética , Prolina Oxidase/metabolismo , Sementes/genética , Sementes/crescimento & desenvolvimento , Sementes/fisiologia
10.
J Biol Chem ; 296: 100660, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33862083

RESUMO

The proteasome is a large protease complex that degrades many different cellular proteins. In eukaryotes, the 26S proteasome contains six different subunits of the ATPases associated with diverse cellular activities family, Rpt1-Rpt6, which form a hexameric ring as part of the base subcomplex that drives unfolding and translocation of substrates into the proteasome core. Archaeal proteasomes contain only a single Rpt-like ATPases associated with diverse cellular activities ATPase, the proteasome-activating nucleotidase, which forms a trimer of dimers. A key proteasome-activating nucleotidase proline residue (P91) forms cis- and trans-peptide bonds in successive subunits around the ring, allowing efficient dimerization through upstream coiled coils. However, the importance of the equivalent Rpt prolines for eukaryotic proteasome assembly was unknown. Here we showed that the equivalent proline is highly conserved in Rpt2, Rpt3, and Rpt5, and loosely conserved in Rpt1, in deeply divergent eukaryotes. Although in no case was a single Pro-to-Ala substitution in budding yeast strongly deleterious to growth, the rpt5-P76A mutation decreased levels of the protein and induced a mild proteasome assembly defect. Moreover, the rpt2-P103A, rpt3-P93A, and rpt5-P76A mutations all caused synthetic defects when combined with deletions of specific proteasome base assembly chaperones. The rpt2-P103A rpt5-P76A double mutant had uniquely strong growth defects attributable to defects in proteasome base formation. Several Rpt subunits in this mutant formed aggregates that were cleared, at least in part, by Hsp42 chaperone-mediated protein quality control. We propose that the conserved Rpt linker prolines promote efficient 26S proteasome base assembly by facilitating specific ATPase heterodimerization.


Assuntos
Proteínas de Choque Térmico/metabolismo , Prolina/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Repressoras/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Sequência de Aminoácidos , Proteínas de Choque Térmico/genética , Mutação , Prolina/genética , Complexo de Endopeptidases do Proteassoma/genética , Ligação Proteica , Domínios Proteicos , Proteínas Repressoras/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Homologia de Sequência , Ubiquitina-Proteína Ligases/genética
11.
Plant Cell Rep ; 40(8): 1429-1450, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-33909122

RESUMO

Heat stress adversely affects plants growth potential. Global warming is reported to increase in the intensity, frequency, and duration of heatwaves, eventually affecting ecology, agriculture and economy. With an expected increase in average temperature by 2-3 °C over the next 30-50 years, crop production is facing a severe threat to sub-optimum growth conditions. Abscisic acid (ABA) and nitric oxide (NO) are growth regulators that are involved in the adaptation to heat stress by affecting each other and changing the adaptation process. The interaction between these molecules has been discussed in various studies in general or under stress conditions; however, regarding high temperature, their interaction has little been worked out. In the present review, the focus is shifted on the role of these molecules under heat stress emphasizing the different possible interactions between ABA and NO as both regulate stomatal closure and other molecules including hydrogen peroxide (H2O2), hydrogen sulfide (H2S), antioxidants, proline, glycine betaine, calcium (Ca2+) and heat shock protein (HSP). Exploring the crosstalk between ABA and NO with other molecules under heat stress will provide us with a comprehensive knowledge of plants mechanism of heat tolerance which could be useful to develop heat stress-resistant varieties.


Assuntos
Ácido Abscísico/metabolismo , Resposta ao Choque Térmico/fisiologia , Óxido Nítrico/metabolismo , Fenômenos Fisiológicos Vegetais , Proteínas de Plantas/metabolismo , Adaptação Fisiológica , Antioxidantes/metabolismo , Betaína/metabolismo , Enzimas/metabolismo , Peróxido de Hidrogênio/metabolismo , Sulfeto de Hidrogênio/metabolismo , Prolina/metabolismo , Transdução de Sinais
12.
Biochem Biophys Res Commun ; 557: 110-116, 2021 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-33862453

RESUMO

The peptide mimicking small extracellular loop of CD82/KAI1 has been reported to inhibit tumor cell migration and metastasis. This provides an evidence that small extracellular loop domain should be important for the function of CD82/KAI1. In this paper, to investigate the structure basis for the function of EC1 mimic peptide, we systematically analyzed the effects of each amino acid residue in EC1 mimic peptide on its bioactivity. We found that the interfering with the folding of secondary structure with proline, a potent breaker of secondary structure, completely abolished the migration and metastasis-inhibitory activity of EC1 mimic peptide. This means that the bioactivity of EC1 mimic peptide was conformation-dependent. Next, we substitute with proline for amino acid residues in the small extracellular ring region of CD82/KAI1 by the site-specific mutations to disrupting secondary structure and detected its effect on the function of CD82/KAI1. The results showed that the disturbing the secondary structure of small extracellular ring completely abolished the migration and metastasis-inhibitory activity of CD82/KAI1. These results further provide direct evidence that the small extracellular ring is an important function region of CD82/KAI1.


Assuntos
Neoplasias da Mama/metabolismo , Proteína Kangai-1/metabolismo , Neoplasias Pulmonares/metabolismo , Animais , Neoplasias da Mama/patologia , Neoplasias da Mama/prevenção & controle , Linhagem Celular Tumoral , Movimento Celular , Feminino , Genes Supressores de Tumor , Humanos , Proteína Kangai-1/química , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Fragmentos de Peptídeos/farmacologia , Prolina/química , Prolina/metabolismo , Domínios Proteicos , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Int J Mol Sci ; 22(6)2021 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-33805821

RESUMO

Drought is the most serious abiotic stress, which significantly reduces crop productivity. The phytohormone ABA plays a pivotal role in regulating stomatal closing upon drought stress. Here, we characterized the physiological function of AtBBD1, which has bifunctional nuclease activity, on drought stress. We found that AtBBD1 localized to the nucleus and cytoplasm, and was expressed strongly in trichomes and stomatal guard cells of leaves, based on promoter:GUS constructs. Expression analyses revealed that AtBBD1 and AtBBD2 are induced early and strongly by ABA and drought, and that AtBBD1 is also strongly responsive to JA. We then compared phenotypes of two AtBBD1-overexpression lines (AtBBD1-OX), single knockout atbbd1, and double knockout atbbd1/atbbd2 plants under drought conditions. We did not observe any phenotypic difference among them under normal growth conditions, while OX lines had greatly enhanced drought tolerance, lower transpirational water loss, and higher proline content than the WT and KOs. Moreover, by measuring seed germination rate and the stomatal aperture after ABA treatment, we found that AtBBD1-OX and atbbd1 plants showed significantly higher and lower ABA-sensitivity, respectively, than the WT. RNA sequencing analysis of AtBBD1-OX and atbbd1 plants under PEG-induced drought stress showed that overexpression of AtBBD1 enhances the expression of key regulatory genes in the ABA-mediated drought signaling cascade, particularly by inducing genes related to ABA biosynthesis, downstream transcription factors, and other regulatory proteins, conferring AtBBD1-OXs with drought tolerance. Taken together, we suggest that AtBBD1 functions as a novel positive regulator of drought responses by enhancing the expression of ABA- and drought stress-responsive genes as well as by increasing proline content.


Assuntos
Ácido Abscísico/metabolismo , Adaptação Fisiológica/genética , Proteínas de Arabidopsis/genética , Arabidopsis/genética , Endonucleases/genética , Regulação da Expressão Gênica de Plantas , Ácido Abscísico/farmacologia , Arabidopsis/efeitos dos fármacos , Arabidopsis/enzimologia , Proteínas de Arabidopsis/agonistas , Proteínas de Arabidopsis/antagonistas & inibidores , Proteínas de Arabidopsis/metabolismo , Núcleo Celular/metabolismo , Ciclopentanos/metabolismo , Ciclopentanos/farmacologia , Citoplasma/metabolismo , Secas , Endonucleases/antagonistas & inibidores , Endonucleases/metabolismo , Isoenzimas/antagonistas & inibidores , Isoenzimas/genética , Isoenzimas/metabolismo , Oxilipinas/metabolismo , Oxilipinas/farmacologia , Células Vegetais/efeitos dos fármacos , Células Vegetais/enzimologia , Reguladores de Crescimento de Plantas/metabolismo , Reguladores de Crescimento de Plantas/farmacologia , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/enzimologia , Folhas de Planta/genética , Estômatos de Plantas/efeitos dos fármacos , Estômatos de Plantas/enzimologia , Estômatos de Plantas/genética , Plantas Geneticamente Modificadas , Prolina/metabolismo , Estresse Fisiológico/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Água/metabolismo
14.
Molecules ; 26(8)2021 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-33920363

RESUMO

As one of the common abiotic stresses, chilling stress has negative effects on rice growth and development. Minimization of these adverse effects through various ways is vital for the productivity of rice. Nanoparticles (NPs) serve as one of the effective alleviation methods against abiotic stresses. In our research, zinc oxide (ZnO) NPs were utilized as foliar sprays on rice leaves to explore the mechanism underlying the effect of NPs against the negative impact of chilling stress on rice seedlings. We revealed that foliar application of ZnO NPs significantly alleviated chilling stress in hydroponically grown rice seedlings, including improved plant height, root length, and dry biomass. Besides, ZnO NPs also restored chlorophyll accumulation and significantly ameliorated chilling-induced oxidative stress with reduced levels of H2O2, MDA, proline, and increased activities of major antioxidative enzymes, superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD). We further found that foliar application of ZnO NPs induced the chilling-induced gene expression of the antioxidative system (OsCu/ZnSOD1, OsCu/ZnSOD2, OsCu/ZnSOD3, OsPRX11, OsPRX65, OsPRX89, OsCATA, and OsCATB) and chilling response transcription factors (OsbZIP52, OsMYB4, OsMYB30, OsNAC5, OsWRKY76, and OsWRKY94) in leaves of chilling-treated seedlings. Taken together, our results suggest that foliar application of ZnO NPs could alleviate chilling stress in rice via the mediation of the antioxidative system and chilling response transcription factors.


Assuntos
Antioxidantes/farmacologia , Clorofila/biossíntese , Nanopartículas/química , Oryza/efeitos dos fármacos , Fatores de Transcrição/genética , Óxido de Zinco/farmacologia , Catalase/genética , Catalase/metabolismo , Clorofila/agonistas , Temperatura Baixa , Regulação da Expressão Gênica de Plantas , Hidroponia/métodos , Malondialdeído/metabolismo , Nanopartículas/ultraestrutura , Oryza/genética , Oryza/crescimento & desenvolvimento , Oryza/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Peroxidase/genética , Peroxidase/metabolismo , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Prolina/metabolismo , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Plântula/efeitos dos fármacos , Plântula/genética , Plântula/crescimento & desenvolvimento , Plântula/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Fatores de Transcrição/agonistas , Fatores de Transcrição/metabolismo
15.
Biochem Biophys Res Commun ; 549: 194-199, 2021 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-33721671

RESUMO

Chemotaxis is the process of sensing chemical gradients and navigating towards favourable conditions. Bacterial chemotaxis is mediated by arrays of trans-membrane chemoreceptor proteins. The most common class of chemoreceptors have periplasmic ligand-binding domains (LBDs) that detect extracellular chemical signs and transduce these signals to the downstream chemotaxis machinery. The repertoire of chemoreceptor proteins in a bacterium determines the range of environmental signals to which it can respond. Pseudomonas syringae pv. actinidiae (Psa) is a plant pathogen which causes bacterial canker of kiwifruit (Actinidia sp.). Compared to many other bacteria, Psa has a large number of chemoreceptors encoded in its genome (43) and most of these remain uncharacterized. A previous study identified PscC as a potential chemoreceptor for l-proline and other amino acid ligands. Here, we have characterized the interaction of PscC-LBD with l-proline using a combination of isothermal titration calorimetry (ITC) and X-ray crystallography. ITC confirmed direct binding of l-proline to PscC-LBD with KD value of 5.0 µM. We determined the structure of PscC-LBD in complex with l-proline. Our structural analysis showed that PscC-LBD adopts similar double-CACHE fold to several other amino acid chemoreceptors. A comparison of the PscC-LDB to other dCACHE structures highlights residues in the binding cavity which contribute to its ligand specificity.


Assuntos
Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Células Quimiorreceptoras/metabolismo , Prolina/metabolismo , Pseudomonas syringae/metabolismo , Sítios de Ligação , Calorimetria , Cristalografia por Raios X , Ligantes , Modelos Moleculares , Domínios Proteicos
16.
Plant Signal Behav ; 16(5): 1899672, 2021 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-33704006

RESUMO

When synchronized with the light/dark cycle the circadian rhythm is termed a diurnal rhythm and this organizes an organism's daily life cycle in relation to the metabolic shifts during the day/night cycles. This is a complex task, particularly under stress conditions. Accurate maintenance of the diurnal rhythm becomes an issue under environmental extremes, such as drought due to the impairment of metabolism, redox balance, and structural integrity. In plants, the non-proteinogenic amino acid GABA accumulates to high levels in response to several stress factors but this is not always dependent on the activation of its biosynthesis. Here we propose a regulatory role to GABA during the diurnal rhythm in plants which is similar to its function in animals where it adjusts the circadian rhythm. Here we investigated whether GABA-biosynthesis was affected by drought stress during the diurnal cycle. For this, we took samples from leaves of N. tabacum plants subjected to PEG-mediated drought stress (-0.73 MPa) during the day and night cycle during a 24 hour period. Glutamate, GABA, and proline contents, along with GDH, GAD enzyme activities and transcript profiles were analyzed. Overall, we conclude that the oscillations in GABA biosynthesis during day and night cycle have an impact on drought stress responses which needs to be elucidated by further analysis.


Assuntos
Ritmo Circadiano/fisiologia , Secas , Tabaco/fisiologia , Ácido gama-Aminobutírico/biossíntese , Glutamato Descarboxilase/metabolismo , Glutamato Desidrogenase/metabolismo , Ácido Glutâmico/metabolismo , Prolina/metabolismo , Tabaco/crescimento & desenvolvimento
17.
Plant Physiol ; 185(2): 352-368, 2021 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-33721894

RESUMO

KLU, encoded by a cytochrome P450 CYP78A family gene, generates an important-albeit unknown-mobile signal that is distinct from the classical phytohormones. Multiple lines of evidence suggest that KLU/KLU-dependent signaling functions in several vital developmental programs, including leaf initiation, leaf/floral organ growth, and megasporocyte cell fate. However, the interactions between KLU/KLU-dependent signaling and the other classical phytohormones, as well as how KLU influences plant physiological responses, remain poorly understood. Here, we applied in-depth, multi-omics analysis to monitor transcriptome and metabolome dynamics in klu-mutant and KLU-overexpressing Arabidopsis plants. By integrating transcriptome sequencing data and primary metabolite profiling alongside phytohormone measurements, our results showed that cytokinin signaling, with its well-established function in delaying leaf senescence, was activated in KLU-overexpressing plants. Consistently, KLU-overexpressing plants exhibited significantly delayed leaf senescence and increased leaf longevity, whereas the klu-mutant plants showed early leaf senescence. In addition, proline biosynthesis and catabolism were enhanced following KLU overexpression owing to increased expression of genes associated with proline metabolism. Furthermore, KLU-overexpressing plants showed enhanced drought-stress tolerance and reduced water loss. Collectively, our work illustrates a role for KLU in positively regulating leaf longevity and drought tolerance by synergistically activating cytokinin signaling and promoting proline metabolism. These data promote KLU as a potential ideal genetic target to improve plant fitness.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Sistema Enzimático do Citocromo P-450/metabolismo , Citocininas/metabolismo , Metaboloma , Reguladores de Crescimento de Plantas/metabolismo , Transdução de Sinais , Transcriptoma , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Sistema Enzimático do Citocromo P-450/genética , Secas , Expressão Gênica , Genômica , Folhas de Planta/enzimologia , Folhas de Planta/genética , Folhas de Planta/fisiologia , Prolina/metabolismo , Estresse Fisiológico
18.
Plant Physiol ; 185(2): 385-404, 2021 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-33721903

RESUMO

During drought stress, cellular proteostasis on the one hand and amino acid homeostasis on the other hand are severely challenged, because the decrease in photosynthesis induces massive proteolysis, leading to drastic changes in both the proteome and the free amino acid pool. Thus, we selected progressive drought stress in Arabidopsis (Arabidopsis thaliana) as a model to investigate on a quantitative level the balance between protein and free amino acid homeostasis. We analyzed the mass composition of the leaf proteome based on proteomics datasets, and estimated how many protein molecules are present in a plant cell and its subcellular compartments. In addition, we calculated stress-induced changes in the distribution of individual amino acids between the free and protein-bound pools. Under control conditions, an average Arabidopsis mesophyll cell contains about 25 billion protein molecules, of which 80% are localized in chloroplasts. Severe water deficiency leads to degradation of more than 40% of the leaf protein mass, and thus causes a drastic shift in distribution toward the free amino acid pool. Stress-induced proteolysis of just half of the 340 million RubisCO hexadecamers present in the chloroplasts of a single mesophyll cell doubles the cellular content of free amino acids. A major fraction of the amino acids released from proteins is channeled into synthesis of proline, which is a compatible osmolyte. Complete oxidation of the remaining fraction as an alternative respiratory substrate can fully compensate for the lack of photosynthesis-derived carbohydrates for several hours.


Assuntos
Aminoácidos/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Proteoma , Cloroplastos/metabolismo , Secas , Homeostase , Fotossíntese , Células Vegetais/fisiologia , Folhas de Planta/fisiologia , Prolina/metabolismo , Proteólise , Proteômica , Ribulose-Bifosfato Carboxilase/metabolismo , Estresse Fisiológico , Água/metabolismo
19.
BMC Plant Biol ; 21(1): 151, 2021 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-33761873

RESUMO

Virola surinamensis is a forest species widely distributed in the estuaries of the Amazon. These ecosystems are susceptible to contamination by Cadmium (Cd), indicating that the plant has strategies for tolerating this metal. The aim of this study was to assess the nitrogen and carbon metabolism of young plants of Ucuúba (Virola surinamensis) in the presence of cadmium with the perspective of the phytoremediation of contaminated environments. The used experimental design was a completely randomized design with five Cd concentrations (0, 15, 30, 45, and 60 mg L- 1), for 60 days. In general, Cd did not affect nitrate concentration in the root but had a positive effect on leaves. The reduction of nitrate reductase (NR) in plants exposed to Cd was followed by a decrease in ammonia, total soluble amino acids (TSA), and total soluble proteins (TSP). Cd promoted an increase in the concentration of total soluble carbohydrates (TSC), proline, sucrose, and reducing sugars in the plants. The increase in TSC, sucrose and proline, suggests a metabolic regulatory mechanism of V. surinamensis against Cd stress.


Assuntos
Cádmio/farmacologia , Myristicaceae/efeitos dos fármacos , Myristicaceae/metabolismo , Poluentes do Solo/farmacologia , Aminoácidos/metabolismo , Compostos de Amônio/metabolismo , Metabolismo dos Carboidratos , Nitrato Redutase/metabolismo , Nitratos/metabolismo , Proteínas de Plantas/metabolismo , Prolina/metabolismo
20.
PLoS Pathog ; 17(3): e1009204, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33647053

RESUMO

Trypanosoma brucei, a protist responsible for human African trypanosomiasis (sleeping sickness), is transmitted by the tsetse fly where the procyclic forms of the parasite develop in the proline-rich (1-2 mM) and glucose-depleted digestive tract. Proline is essential for the midgut colonization of the parasite in the insect vector, however other carbon sources could be available and used to feed its central metabolism. Here we show that procyclic trypanosomes can consume and metabolize metabolic intermediates, including those excreted from glucose catabolism (succinate, alanine and pyruvate), with the exception of acetate, which is the ultimate end-product excreted by the parasite. Among the tested metabolites, tricarboxylic acid (TCA) cycle intermediates (succinate, malate and α-ketoglutarate) stimulated growth of the parasite in the presence of 2 mM proline. The pathways used for their metabolism were mapped by proton-NMR metabolic profiling and phenotypic analyses of thirteen RNAi and/or null mutants affecting central carbon metabolism. We showed that (i) malate is converted to succinate by both the reducing and oxidative branches of the TCA cycle, which demonstrates that procyclic trypanosomes can use the full TCA cycle, (ii) the enormous rate of α-ketoglutarate consumption (15-times higher than glucose) is possible thanks to the balanced production and consumption of NADH at the substrate level and (iii) α-ketoglutarate is toxic for trypanosomes if not appropriately metabolized as observed for an α-ketoglutarate dehydrogenase null mutant. In addition, epimastigotes produced from procyclics upon overexpression of RBP6 showed a growth defect in the presence of 2 mM proline, which is rescued by α-ketoglutarate, suggesting that physiological amounts of proline are not sufficient per se for the development of trypanosomes in the fly. In conclusion, these data show that trypanosomes can metabolize multiple metabolites, in addition to proline, which allows them to confront challenging environments in the fly.


Assuntos
Glucose/metabolismo , Prolina/farmacologia , Trypanosoma brucei brucei/efeitos dos fármacos , Trypanosoma/efeitos dos fármacos , Moscas Tsé-Tsé/efeitos dos fármacos , Animais , Ciclo do Ácido Cítrico/efeitos dos fármacos , Insetos Vetores/parasitologia , Oxirredução/efeitos dos fármacos , Prolina/metabolismo , Interferência de RNA/fisiologia , Trypanosoma/metabolismo , Trypanosoma brucei brucei/metabolismo , Tripanossomíase Africana/tratamento farmacológico , Moscas Tsé-Tsé/parasitologia
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