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1.
Curr Microbiol ; 79(6): 181, 2022 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-35508788

RESUMO

Vibrio fluvialis is a marine opportunistic pathogen that frequently causes diseases in aquatic animals and humans. V. fluvialis can produce quorum sensing signaling molecules to coordinate cell density-dependent behavioral changes, including N-acyl homoserine lactone (AHL), which acts as a vital mediator of virulence-associated gene expression. Currently, several AHL molecules in V. fluvialis have been detected via biological and physicochemical methods, although different detection approaches have generated diverse AHL profiles. Here, we describe the AHL-producing bacterium, V. fluvialis BJ-1, which was isolated from marine sediments from the East China Sea. V. fluvialis BJ-1 could stimulate AHL-mediated ß-galactosidase synthesis of the biosensor Agrobacterium tumefaciens NTL4 (pZLR4) but could not induce violacein production in the AHL reporter strain, Chromobacterium violaceum CV026. This bacterial isolate exhibited strong AHL-producing activity at low cell density; however, the AHL activity declined when population density remained at high levels. Analysis of the AHLs by Ultra-High-Performance Liquid Chromatography tandem Mass Spectrometry demonstrated that V. fluvialis BJ-1 produced five different AHL signaling molecules, including two linear chain AHL products (C8- and C10-HSL), and three ß-carbon-oxidative AHL products (3-O-C8-, 3-O-C10- and 3-O-C12-HSL). Significantly, the present study is the first to accurately define the AHL profile of marine V. fluvialis. In future, the coupling of UHPLC to ESI-MS/MS is expected to be utilized for the accurate determination of AHL profiles in marine Vibrio.


Assuntos
Acil-Butirolactonas , Vibrio , 4-Butirolactona/metabolismo , Animais , Cromatografia Líquida de Alta Pressão , Homosserina/metabolismo , Lactonas/metabolismo , Percepção de Quorum , Espectrometria de Massas em Tandem , Vibrio/genética , Vibrio/metabolismo
2.
Plant Commun ; 3(2): 100303, 2022 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-35529949

RESUMO

The effects of the phytohormone strigolactone (SL) and smoke-derived karrikins (KARs) on plants are generally distinct, despite the fact that they are perceived through very similar mechanisms. The homologous receptors DWARF14 (D14) and KARRIKIN-INSENSITIVE2 (KAI2), together with the F-box protein MORE AXILLARY GROWTH2 (MAX2), mediate SL and KAR responses, respectively, by targeting different SMAX1-LIKE (SMXL) family proteins for degradation. These mechanisms are putatively well-insulated, with D14-MAX2 targeting SMXL6, SMXL7, and SMXL8 and KAI2-MAX2 targeting SMAX1 and SMXL2 in Arabidopsis thaliana. Recent evidence challenges this model. We investigated whether D14 can target SMAX1 and whether this occurs naturally. Genetic analysis indicates that the SL analog GR24 promotes D14-SMAX1 crosstalk. Although D14 shows weaker interactions with SMAX1 than with SMXL2 or SMXL7, D14 mediates GR24-induced degradation of SMAX1 in plants. Osmotic stress triggers SMAX1 degradation, which is protective, through SL biosynthesis and signaling genes. Thus, D14-SMAX1 crosstalk may be beneficial and not simply a vestige of the evolution of the SL pathway.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Proteínas de Transporte/metabolismo , Proteínas de Transporte/farmacologia , Compostos Heterocíclicos com 3 Anéis , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lactonas/metabolismo , Lactonas/farmacologia , Pressão Osmótica
3.
Physiol Plant ; 174(2): e13681, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35362177

RESUMO

Strigolactones (SLs) are the most recently discovered phytohormones, and their roles in root architecture and metabolism are not fully understood. Here, we investigated four MORE AXILLARY GROWTH (MAX) SL mutants in Arabidopsis thaliana, max3-9, max4-1, max1-1 and max2-1, as well as the SL receptor mutant d14-1 and karrikin receptor mutant kai2-2. By characterising max2-1 and max4-1, we found that variation in SL biosynthesis modified multiple metabolic pathways in root tissue, including that of xyloglucan, triterpenoids, fatty acids and flavonoids. The transcription of key flavonoid biosynthetic genes, including TRANSPARENT TESTA4 (TT4) and TRANSPARENT TESTA5 (TT5) was downregulated in max2 roots and seedlings, indicating that the proposed MAX2 regulation of flavonoid biosynthesis has a widespread effect. We found an enrichment of BRI1-EMS-SUPPRESSOR 1 (BES1) targets amongst genes specifically altered in the max2 mutant, reflecting that the regulation of flavonoid biosynthesis likely occurs through the MAX2 degradation of BES1, a key brassinosteroid-related transcription factor. Finally, flavonoid accumulation decreased in max2-1 roots, supporting a role for MAX2 in regulating both SL and flavonoid biosynthesis.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Flavonoides/metabolismo , Regulação da Expressão Gênica de Plantas , Compostos Heterocíclicos com 3 Anéis , Lactonas/metabolismo
4.
J Phys Chem B ; 126(11): 2188-2195, 2022 03 24.
Artigo em Inglês | MEDLINE | ID: mdl-35275626

RESUMO

Plant branch and root growth relies on metabolism of the strigolactone (SL) hormone. The interaction between the SL molecule, Oryza sativa DWARF14 (D14) SL receptor, and D3 F-box protein has been shown to play a critical role in SL perception. Previously, it was believed that D3 only interacts with the closed form of D14 to induce downstream signaling, but recent experiments indicate that D3, as well as its C-terminal helix (CTH), can interact with the open form as well to inhibit strigolactone signaling. Two hypotheses for the CTH induced inhibition are that either the CTH affects the conformational ensemble of D14 by stabilizing catalytically inactive states or the CTH interacts with SLs in a way that prevents them from entering the binding pocket. In this study, we have performed molecular dynamics (MD) simulations to assess the validity of these hypotheses. We used an apo system with only D14 and the CTH to test the active site conformational stability and a holo system with D14, the CTH, and an SL molecule to test the interaction between the SL and CTH. Our simulations show that the CTH affects both active site conformation and the ability of SLs to move into the binding pocket. In the apo system, the CTH allosterically stabilized catalytic residues into their inactive conformation. In the holo system, significant interactions between SLs and the CTH hindered the ability of SLs to enter the D14 binding pocket. These two mechanisms account for the observed decrease in SL binding to D14 and subsequent ligand hydrolysis in the presence of the CTH.


Assuntos
Oryza , Reguladores de Crescimento de Plantas , Compostos Heterocíclicos com 3 Anéis , Hidrólise , Lactonas/química , Lactonas/metabolismo , Oryza/metabolismo , Reguladores de Crescimento de Plantas/química , Reguladores de Crescimento de Plantas/metabolismo , Proteínas de Plantas/química
5.
Proc Natl Acad Sci U S A ; 119(14): e2111565119, 2022 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-35344437

RESUMO

SignificanceStrigolactones (SLs) are a group of apocarotenoid hormones, which regulates shoot branching and other diverse developmental processes in plants. The major bioactive form(s) of SLs as endogenous hormones has not yet been clarified. Here, we identify an Arabidopsis methyltransferase, CLAMT, responsible for the conversion of an inactive precursor to a biologically active SL that can interact with the SL receptor in vitro. Reverse genetic analysis showed that this enzyme plays an essential role in inhibiting shoot branching. This mutant also contributed to specifying the SL-related metabolites that could move from root to shoot in grafting experiments. Our work has identified a key enzyme necessary for the production of the bioactive form(s) of SLs.


Assuntos
Arabidopsis , Arabidopsis/genética , Arabidopsis/metabolismo , Hormônios/metabolismo , Lactonas/metabolismo , Metiltransferases/genética , Metiltransferases/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Brotos de Planta/genética , Brotos de Planta/metabolismo
6.
J Chem Inf Model ; 62(7): 1712-1722, 2022 04 11.
Artigo em Inglês | MEDLINE | ID: mdl-35192364

RESUMO

Parasitic weeds such as Striga have led to significant losses in agricultural productivity worldwide. These weeds use the plant hormone strigolactone as a germination stimulant. Strigolactone signaling involves substrate hydrolysis followed by a conformational change of the receptor to a "closed" or "active" state that associates with a signaling partner, MAX2/D3. Crystal structures of active and inactive AtD14 receptors have helped elucidate the structural changes involved in activation. However, the mechanism by which the receptor activates remains unknown. The ligand dependence of AtD14 activation has been disputed by mutagenesis studies showing that enzymatically inactive receptors are able to associate with MAX2 proteins. Furthermore, activation differences between strigolactone receptor in Striga, ShHTL7, and AtD14 could contribute to the high sensitivity to strigolactones exhibited by parasitic plants. Using molecular dynamics simulations, we demonstrate that both AtD14 and ShHTL7 could adopt an active conformation in the absence of ligand. However, ShHTL7 exhibits a higher population in the inactive apo state as compared to the AtD14 receptor. We demonstrate that this difference in inactive state population is caused by sequence differences between their D-loops and interactions with the catalytic histidine that prevent full binding pocket closure in ShHTL7. These results indicate that ligand hydrolysis would enhance the active state population by destabilizing the inactive state in ShHTL7 as compared to AtD14. We also show that the mechanism of activation is more concerted in AtD14 than in ShHTL7 and that the main barrier to activation in ShHTL7 is closing of the binding pocket.


Assuntos
Striga , Proteínas de Transporte/metabolismo , Compostos Heterocíclicos com 3 Anéis/química , Compostos Heterocíclicos com 3 Anéis/metabolismo , Lactonas/química , Lactonas/metabolismo , Lactonas/farmacologia , Ligantes , Plantas Daninhas/química , Plantas Daninhas/metabolismo , Striga/química , Striga/metabolismo
7.
New Phytol ; 234(3): 1003-1017, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35119708

RESUMO

Strigolactones (SLs) are plant hormones and important signalling molecules required to promote arbuscular mycorrhizal (AM) symbiosis. While in plants an α/ß-hydrolase, DWARF14 (D14), was shown to act as a receptor that binds and cleaves SLs, the fungal receptor for SLs is unknown. Since AM fungi are currently not genetically tractable, in this study, we used the fungal pathogen Cryphonectria parasitica, for which gene deletion protocols exist, as a model, as we have previously shown that it responds to SLs. By means of computational, biochemical and genetic analyses, we identified a D14 structural homologue, CpD14. Molecular homology modelling and docking support the prediction that CpD14 interacts with and hydrolyses SLs. The recombinant CpD14 protein shows α/ß hydrolytic activity in vitro against the SLs synthetic analogue GR24; its enzymatic activity requires an intact Ser/His/Asp catalytic triad. CpD14 expression in the d14-1 loss-of-function Arabidopsis thaliana line did not rescue the plant mutant phenotype. However, gene inactivation by knockout homologous recombination reduced fungal sensitivity to SLs. These results indicate that CpD14 is involved in SLs responses in C. parasitica and strengthen the role of SLs as multifunctional molecules acting in plant-microbe interactions.


Assuntos
Ascomicetos , Proteínas de Plantas , Ascomicetos/metabolismo , Compostos Heterocíclicos com 3 Anéis , Lactonas/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Proteínas de Plantas/metabolismo
8.
Toxicol Lett ; 358: 88-99, 2022 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-35104616

RESUMO

Several Alternaria mycotoxins are believed to act as endocrine disruptive chemicals (EDCs), since they are reported to bind estrogen receptors in several experimental models. After ingestion of contaminated food commodities, the mycotoxins reach the intestine, where they come into direct contact with food constituents as well as the gut microbiota. Thus, the aim of the present work was to evaluate the modulatory potential of a complex extract of cultured Alternaria fungi (CE; containing eleven chemically characterized compounds) on the estrogenic signaling cascade of mammalian cells before and after anaerobic incubation with fecal slurries, in order to simulate an in vivo-like condition in the gut. Assessing alkaline phosphatase expression in Ishikawa cells as a measure for estrogenicity, we found the CE to partially quench the intrinsic estrogenic properties of fecal slurries and fecal waters, even after 3 h of fecal incubation. Investigation of the mechanisms underlying the effects observed carried out through an in vitro/in silico approach revealed the ability of the extract to decrease the ERα/ERß nuclear ratio, while a possible action of the mycotoxins as ER-antagonists was excluded. Our results suggest that Alternaria mycotoxins might act as EDCs in vivo, and warrant further investigation in animal models.


Assuntos
Micotoxinas , Alternaria/metabolismo , Anaerobiose , Animais , Estrogênios/metabolismo , Fezes/química , Contaminação de Alimentos/análise , Humanos , Lactonas/metabolismo , Mamíferos/metabolismo , Micotoxinas/metabolismo
9.
J Plant Res ; 135(2): 337-350, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35106650

RESUMO

Both strigolactones (SLs) and nitric oxide (NO) are regulatory signals with diverse roles during stress responses. At present, the interaction and mechanism of SLs and NO in tomato salt tolerance remain unclear. In the current study, tomato 'Micro-Tom' was used to study the roles and interactions of SLs and NO in salinity stress tolerance. The results show that 15 µM SLs synthetic analogs GR24 and 10 µM NO donor S-nitrosoglutathione (GSNO) promoted seedling growth under salt stress. TIS108 (an inhibitor of strigolactone synthesis) suppressed the positive roles of NO in tomato growth under salt stress, indicating that endogenous SLs might be involved in NO-induced salt response in tomato seedlings. Meanwhile, under salt stress, GSNO or GR24 treatment induced the increase of endogenous SLs content in tomato seedlings. Moreover, GR24 or GSNO treatment effectively increased the content of chlorophyll, carotenoids and ascorbic acid (ASA), and enhanced the activities of antioxidant enzymes (superoxide dismutase, peroxidase, catalase, and ascorbate peroxidase), glutathione reductase (GR) and cleavage dioxygenase (CCD) enzyme. Additionally, GSNO or GR24 treatment also up-regulated the expression of SLs synthesis genes (SlCCD7, SlCCD8, SlD27 and SlMAX1) and its signal transduction genes (SlD14 and SlMAX2) in tomato seedlings under salt stress. While, a strigolactone synthesis inhibitor TIS108 blocked the increase of endogenous SLs, chlorophyll, carotenoids and ASA content, and antioxidant enzyme, GR, CCD enzyme activity and SLs-related gene expression levels induced by GSNO. Thus, SLs may play an important role in NO-enhanced salinity tolerance in tomato seedlings by increasing photosynthetic pigment content, enhancing antioxidant capacity and improving endogenous SLs synthesis.


Assuntos
Lycopersicon esculentum , Plântula , Antioxidantes/metabolismo , Compostos Heterocíclicos com 3 Anéis , Lactonas/metabolismo , Lactonas/farmacologia , Óxido Nítrico/metabolismo , Plântula/fisiologia
10.
BMC Plant Biol ; 22(1): 30, 2022 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-35027005

RESUMO

Strigolactone is a newly discovered type of plant hormone that has multiple roles in modulating plant responses to abiotic stress. Herein, we aimed to investigate the effects of exogenous GR24 (a synthetic analogue of strigolactone) on plant growth, photosynthetic characteristics, carbohydrate levels, endogenous strigolactone content and antioxidant metabolism in cucumber seedlings under low light stress. The results showed that the application of 10 µM GR24 can increase the photosynthetic efficiency and plant biomass of low light-stressed cucumber seedlings. GR24 increased the accumulation of carbohydrates and the synthesis of sucrose-related enzyme activities, enhanced antioxidant enzyme activities and antioxidant substance contents, and reduced the levels of H2O2 and MDA in cucumber seedlings under low light stress. These results indicate that exogenous GR24 might alleviate low light stress-induced growth inhibition by regulating the assimilation of carbon and antioxidants and endogenous strigolactone contents, thereby enhancing the tolerance of cucumber seedlings to low light stress.


Assuntos
Adaptação Ocular/efeitos dos fármacos , Cucumis sativus/efeitos dos fármacos , Cucumis sativus/crescimento & desenvolvimento , Cucumis sativus/metabolismo , Compostos Heterocíclicos com 3 Anéis/metabolismo , Lactonas/metabolismo , Produtos Agrícolas/efeitos dos fármacos , Produtos Agrícolas/crescimento & desenvolvimento , Produtos Agrícolas/metabolismo
11.
Sci Rep ; 12(1): 203, 2022 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-34997073

RESUMO

Burkholderia pseudomallei is a saprophytic bacterium endemic throughout the tropics causing severe disease in humans and animals. Environmental signals such as the accumulation of inorganic ions mediates the biofilm forming capabilities and survival of B. pseudomallei. We have previously shown that B. pseudomallei responds to nitrate and nitrite by inhibiting biofilm formation and altering cyclic di-GMP signaling. To better understand the roles of nitrate-sensing in the biofilm inhibitory phenotype of B. pseudomallei, we created in-frame deletions of narX (Bp1026b_I1014) and narL (Bp1026b_I1013), which are adjacent components of a conserved nitrate-sensing two-component system. We observed transcriptional downregulation in key components of the biofilm matrix in response to nitrate and nitrite. Some of the most differentially expressed genes were nonribosomal peptide synthases (NRPS) and/or polyketide synthases (PKS) encoding the proteins for the biosynthesis of bactobolin, malleilactone, and syrbactin, and an uncharacterized cryptic NRPS biosynthetic cluster. RNA expression patterns were reversed in ∆narX and ∆narL mutants, suggesting that nitrate sensing is an important checkpoint for regulating the diverse metabolic changes occurring in the biofilm inhibitory phenotype. Moreover, in a macrophage model of infection, ∆narX and ∆narL mutants were attenuated in intracellular replication, suggesting that nitrate sensing contributes to survival in the host.


Assuntos
Proteínas de Bactérias/metabolismo , Biofilmes/crescimento & desenvolvimento , Produtos Biológicos/metabolismo , Burkholderia pseudomallei/metabolismo , Nitratos/metabolismo , Nitritos/metabolismo , Proteínas de Bactérias/genética , Benzopiranos/metabolismo , Burkholderia pseudomallei/genética , Burkholderia pseudomallei/crescimento & desenvolvimento , Regulação Bacteriana da Expressão Gênica , Interações Hospedeiro-Patógeno , Lactonas/metabolismo , Viabilidade Microbiana , Mutação , Transcrição Genética
12.
Sci Rep ; 12(1): 831, 2022 01 17.
Artigo em Inglês | MEDLINE | ID: mdl-35039556

RESUMO

Roles for the non-coding small RNA RyhB in quorum-sensing and iron-dependent gene modulation in the human pathogen V. vulnificus were assessed in this study. Both the quorum sensing master regulator SmcR and the Fur-iron complex were observed to bind to the region upstream of the non-coding small RNA RyhB gene to repress expression, which suggests that RyhB is associated with both quorum-sensing and iron-dependent signaling in this pathogen. We found that expression of LuxS, which is responsible for the biosynthesis of autoinducer-2 (AI-2), was higher in wild type than in a ryhB-deletion isotype. RyhB binds directly to the 5'-UTR (untranslated region) of the luxS transcript to form a heteroduplex, which not only stabilizes luxS mRNA but also disrupts the secondary structure that normally obscures the translational start codon and thereby allows translation of LuxS to begin. The binding of RyhB to luxS mRNA requires the chaperone protein Hfq, which stabilizes RyhB. These results demonstrate that the small RNA RyhB is a key element associated with feedback control of AI-2 production, and that it inhibits quorum-sensing signaling in an iron-dependent manner. This study, taken together with previous studies, shows that iron availability and cell density signals are funneled to SmcR and RyhB, and that these regulators coordinate cognate signal pathways that result in the proper balance of protein expression in response to environmental conditions.


Assuntos
Genes Bacterianos/genética , Homosserina/análogos & derivados , Ferro/metabolismo , Lactonas/metabolismo , Percepção de Quorum/fisiologia , Pequeno RNA não Traduzido/genética , Pequeno RNA não Traduzido/fisiologia , Vibrio vulnificus/genética , Vibrio vulnificus/fisiologia , Regiões 5' não Traduzidas , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/fisiologia , Liases de Carbono-Enxofre/genética , Liases de Carbono-Enxofre/metabolismo , Liases de Carbono-Enxofre/fisiologia , Regulação Bacteriana da Expressão Gênica/genética , Homosserina/biossíntese , Homosserina/metabolismo , RNA Mensageiro , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Vibrio vulnificus/metabolismo
13.
ACS Synth Biol ; 11(2): 747-759, 2022 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-35090122

RESUMO

Bacteria produce and react to interspecies signaling molecules in order to control the expression of genes that are particularly beneficial when they are expressed by a bacterial community. In addition to intraspecies communication, the signaling molecule autoinducer-2 (AI-2) can also serve for interspecies communication between Gram-positive and Gram-negative bacteria and is therefore of particular interest. The analysis and quantification of AI-2 are essential for understanding population density-dependent changes in bacterial behavior and pathogenicity. However, currently available bioassays for AI-2 quantification are rather complex, have narrow detection ranges, and are very sensitive to trace components of, for example, growth media. To facilitate and improve the detection of AI-2, we have developed an Escherichia coli biosensor-based assay that is sensitive, cheap, fast, robust, and reliable in the quantification of biologically active AI-2. The bioassay is based on an lsr promoter-fluorescent reporter gene fusion cassette that we chromosomally integrated in a biosensor strain, but the cassette can also be used in a low-copy number plasmid for the application in other Gram-negative bacterial species. We show here that AI-2 quantification was possible in a concentration range from 400 nM to 100 µM and that a critical interpretation of the kinetics of the measurements can reveal sugar interference. With the help of our biosensor strain, coculture experiments were done to test the capability and kinetics of AI-2 secretion by various Gram-negative bacteria in real time. Finally, calibration curves were used to calculate the absolute AI-2 concentration in cell-free bacterial samples.


Assuntos
Antibacterianos , Técnicas Biossensoriais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Bioensaio , Bactérias Gram-Negativas/metabolismo , Bactérias Gram-Positivas , Homosserina/metabolismo , Lactonas/metabolismo , Percepção de Quorum
14.
New Phytol ; 234(1): 122-136, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-34716593

RESUMO

Shoot branching is regulated by multiple signals. Previous studies have indicated that sucrose may promote shoot branching through suppressing the inhibitory effect of the hormone strigolactone (SL). However, the molecular mechanisms underlying this effect are unknown. Here, we used molecular and genetic tools to identify the molecular targets underlying the antagonistic interaction between sucrose and SL. We showed that sucrose antagonizes the suppressive action of SL on tillering in rice and on the degradation of D53, a major target of SL signalling. Sucrose inhibits the gene expression of D3, the orthologue of the Arabidopsis F-box MAX2 required for SL signalling. Overexpression of D3 antagonizes sucrose inhibition of D53 degradation and enables the SL inhibition of tillering under high sucrose. Sucrose prevents SL-induced degradation of D14, the SL receptor involved in D53 degradation. In contrast to D3, D14 overexpression enhances D53 protein levels and sucrose-induced tillering, even in the presence of SL. Our results show that sucrose inhibits SL response by affecting key components of SL signalling and, together with previous studies reporting the inhibition of SL synthesis by nitrate and phosphate, demonstrate the central role played by SLs in the regulation of plant architecture by nutrients.


Assuntos
Arabidopsis , Oryza , Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , Lactonas/metabolismo , Lactonas/farmacologia , Oryza/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Sacarose/metabolismo , Sacarose/farmacologia
15.
Plant Physiol ; 188(1): 97-110, 2022 01 20.
Artigo em Inglês | MEDLINE | ID: mdl-34718781

RESUMO

Strigolactones (SLs) are a class of plant hormones that mediate biotic interactions and modulate developmental programs in response to endogenous and exogenous stimuli. However, a comprehensive view on the spatio-temporal pattern of SL signaling has not been established, and tools for a systematic in planta analysis do not exist. Here, we present Strigo-D2, a genetically encoded ratiometric SL signaling sensor that enables the examination of SL signaling distribution at cellular resolution and is capable of rapid response to altered SL levels in intact Arabidopsis (Arabidopsis thaliana) plants. By monitoring the abundance of a truncated and fluorescently labeled SUPPRESSOR OF MAX2 1-LIKE 6 (SMXL6) protein, a proteolytic target of the SL signaling machinery, we show that all cell types investigated have the capacity to respond to changes in SL levels but with very different dynamics. In particular, SL signaling is pronounced in vascular cells but low in guard cells and the meristematic region of the root. We also show that other hormones leave Strigo-D2 activity unchanged, indicating that initial SL signaling steps work in isolation from other hormonal signaling pathways. The specificity and spatio-temporal resolution of Strigo-D2 underline the value of the sensor for monitoring SL signaling in a broad range of biological contexts with highly instructive analytical depth.


Assuntos
Arabidopsis/crescimento & desenvolvimento , Arabidopsis/genética , Arabidopsis/metabolismo , Compostos Heterocíclicos com 3 Anéis/metabolismo , Lactonas/metabolismo , Meristema/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Transdução de Sinais/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Meristema/genética , Meristema/crescimento & desenvolvimento , Reguladores de Crescimento de Plantas/genética , Transdução de Sinais/genética
16.
Protein Sci ; 31(2): 371-383, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34761460

RESUMO

Xylonolactonase Cc XylC from Caulobacter crescentus catalyzes the hydrolysis of the intramolecular ester bond of d-xylonolactone. We have determined crystal structures of Cc XylC in complex with d-xylonolactone isomer analogues d-xylopyranose and (r)-(+)-4-hydroxy-2-pyrrolidinone at high resolution. Cc XylC has a 6-bladed ß-propeller architecture, which contains a central open channel having the active site at one end. According to our previous native mass spectrometry studies, Cc XylC is able to specifically bind Fe2+ . The crystal structures, presented here, revealed an active site bound metal ion with an octahedral binding geometry. The side chains of three amino acid residues, Glu18, Asn146, and Asp196, which participate in binding of metal ion are located in the same plane. The solved complex structures allowed suggesting a reaction mechanism for intramolecular ester bond hydrolysis in which the major contribution for catalysis arises from the carbonyl oxygen coordination of the xylonolactone substrate to the Fe2+ . The structure of Cc XylC was compared with eight other ester hydrolases of the ß-propeller hydrolase family. The previously published crystal structures of other ß-propeller hydrolases contain either Ca2+ , Mg2+ , or Zn2+ and show clear similarities in ligand and metal ion binding geometries to that of Cc XylC. It would be interesting to reinvestigate the metal binding specificity of these enzymes and clarify whether they are also able to use Fe2+ as a catalytic metal. This could further expand our understanding of utilization of Fe2+ not only in oxidative enzymes but also in hydrolases.


Assuntos
Hidrolases de Éster Carboxílico , Caulobacter crescentus , Hidrolases de Éster Carboxílico/química , Hidrolases de Éster Carboxílico/metabolismo , Domínio Catalítico , Caulobacter crescentus/enzimologia , Cristalografia por Raios X , Hidrolases , Hidrólise , Ferro , Lactonas/química , Lactonas/metabolismo
17.
J Exp Bot ; 73(7): 1992-2004, 2022 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-34850875

RESUMO

Root parasitic weeds of the Orobanchaceae, such as witchweeds (Striga spp.) and broomrapes (Orobanche and Phelipanche spp.), cause serious losses in agriculture worldwide, and efforts have been made to control these parasitic weeds. Understanding the characteristic physiological processes in the life cycle of root parasitic weeds is particularly important to identify specific targets for growth modulators. In our previous study, planteose metabolism was revealed to be activated soon after the perception of strigolactones in germinating seeds of O. minor. Nojirimycin inhibited planteose metabolism and impeded seed germination of O. minor, indicating a possible target for root parasitic weed control. In the present study, we investigated the distribution of planteose in dry seeds of O. minor by matrix-assisted laser desorption/ionization-mass spectrometry imaging. Planteose was detected in tissues surrounding-but not within-the embryo, supporting its suggested role as a storage carbohydrate. Biochemical assays and molecular characterization of an α-galactosidase family member, OmAGAL2, indicated that the enzyme is involved in planteose hydrolysis in the apoplast around the embryo after the perception of strigolactones, to provide the embryo with essential hexoses for germination. These results indicate that OmAGAL2 is a potential molecular target for root parasitic weed control.


Assuntos
Orobanche , Germinação/fisiologia , Hidrólise , Lactonas/metabolismo , Raízes de Plantas/metabolismo , Plantas Daninhas/metabolismo , Sementes , alfa-Galactosidase
18.
Plant Physiol ; 188(1): 255-267, 2022 01 20.
Artigo em Inglês | MEDLINE | ID: mdl-34687296

RESUMO

During secondary growth, meristematic cells in the cambium can either proliferate to maintain the stem cell population or differentiate into xylem or phloem. The balance between these two developmental trajectories is tightly regulated by many environmental and endogenous cues. Strigolactones (SLs), a class of plant hormones, were previously reported to regulate secondary growth by promoting cambium activity. However, the underlying molecular mechanisms of SL action in plant secondary growth are not well understood. We performed histological, genetic, and biochemical analyses using genetic materials in Arabidopsis (Arabidopsis thaliana) with altered activity of the transcription factors BRI1-EMS-SUPPRESSOR1 (BES1) or WUSCHEL-related HOMEOBOX4 (WOX4) or lacking MORE AXILLARY SHOOT2 (MAX2), a key positive component in the SL signaling pathway. We found that BES1, a downstream regulator in the SL signaling pathway that promotes shoot branching and xylem differentiation, also inhibits WOX4 expression, a key regulator of cambium cell division in the intercellular TRACHEARY ELEMENT DIFFERENTIATION INHIBITORY FACTOR (TDIF)-TDIF RECEPTOR (TDR) signaling pathway. The antagonistic roles of BES1 and WOX4 in the regulation of cambium activity may integrate intercellular TDIF signals to efficiently and bidirectionally modulate cambium cell proliferation and differentiation. As both BES1 and WOX4 are widely involved in various endogenous signals and responses to environmental stimuli, these findings may provide insight into the dynamic regulation of cambium development.


Assuntos
Arabidopsis/genética , Arabidopsis/metabolismo , Câmbio/metabolismo , Compostos Heterocíclicos com 3 Anéis/metabolismo , Proteínas de Homeodomínio/metabolismo , Lactonas/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fatores de Transcrição , Câmbio/genética , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Variação Genética , Genótipo , Proteínas de Homeodomínio/genética
19.
New Phytol ; 233(4): 1900-1914, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34839530

RESUMO

Light quality affects mutualisms between plant roots and arbuscular mycorrhizal fungi (AMFs), which modify nutrient acquisition in plants. However, the mechanisms by which light systemically modulates root colonization by AMFs and phosphate uptake in roots remain unclear. We used a range of approaches, including grafting techniques, protein immunoblot analysis, electrophoretic mobility shift assay, chromatin immunoprecipitation, and dual-luciferase assays, to unveil the molecular basis of light signal transmission from shoot to root that mediates arbuscule development and phosphate uptake in tomato. The results show that shoot phytochrome B (phyB) triggers shoot-derived mobile ELONGATED HYPOCOTYL5 (HY5) protein accumulation in roots, and HY5 further positively regulates transcription of strigolactone (SL) synthetic genes, thus forming a shoot phyB-dependent systemic signaling pathway that regulates the synthesis and accumulation of SLs in roots. Further experiments with carotenoid cleavage dioxygenase 7 mutants and supplementary red light confirm that SLs are indispensable in the red-light-regulated mycorrhizal symbiosis in roots. Our results reveal a phyB-HY5-SLs systemic signaling cascade that facilitates mycorrhizal symbiosis and phosphate utilization in plants. The findings provide new prospects for the potential application of AMFs and light manipulation to effectively improve nutrient utilization and minimize the use of chemical fertilizers and associated pollution.


Assuntos
Lycopersicon esculentum , Micorrizas , Compostos Heterocíclicos com 3 Anéis , Lactonas/metabolismo , Lycopersicon esculentum/genética , Micorrizas/fisiologia , Raízes de Plantas/metabolismo , Simbiose
20.
Plant Cell Environ ; 45(2): 512-527, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34719040

RESUMO

Nitrogen (N) and phosphorus (P) are among the most important macronutrients for plant growth and development, and the most widely used as fertilizers. Understanding how plants sense and respond to N and P deficiency is essential to optimize and reduce the use of chemical fertilizers. Strigolactones (SLs) are phytohormones acting as modulators and sensors of plant responses to P deficiency. In the present work, we assess the potential role of SLs in N starvation and in the N-P signalling interplay. Physiological, transcriptional and metabolic responses were analysed in wild-type and SL-deficient tomato plants grown under different P and N regimes, and in plants treated with a short-term pulse of the synthetic SL analogue 2'-epi-GR24. The results evidence that plants prioritize N over P status by affecting SL biosynthesis. We also show that SLs modulate the expression of key regulatory genes of phosphate and nitrate signalling pathways, including the N-P integrators PHO2 and NIGT1/HHO. The results support a key role for SLs as sensors during early plant responses to both N and phosphate starvation and mediating the N-P signalling interplay, indicating that SLs are involved in more physiological processes than so far proposed.


Assuntos
Compostos Heterocíclicos com 3 Anéis/metabolismo , Lactonas/metabolismo , Lycopersicon esculentum/fisiologia , Nitrogênio/fisiologia , Fósforo/fisiologia , Transdução de Sinais
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