RESUMO
In the investigation of heterotrimeric G protein-mediated signal transduction in planta, their roles in the transmittance of low K+ stimuli remain to be elucidated. Here, we found that the primary root growth of wild-type Arabidopsis was gradually inhibited with the decrease of external K+ concentrations, while the primary root of the mutants for G protein ß subunit AGB1 and γ subunits AGG1, AGG2 and AGG3 could still grow under low K+ conditions (LK). Exogenous NAA application attenuated primary root elongation in agb1 and agg1/2/3 but promoted the growth in wild-type seedlings under LK stress. Using ProDR5:GFP, ProPIN1:PIN1-GFP and ProPIN2:PIN2-GFP reporter lines, a diminishment in auxin concentration at the radicle apex and a reduction in PIN1and PIN2 efflux carrier abundance were observed in wild-type roots under LK, a phenomenon not recorded in the agb1 and agg1/2/3. Further proteolytic and transcriptional assessments revealed an enhanced degradation of PIN1 and a suppressed expression of PIN2 in the wild-type background under LK, contrasting with the stability observed in the agb1 and agg1/2/3 mutants. Our results indicate that the G protein ß and γ subunits play pivotal roles in suppressing of Arabidopsis root growth under LK by modulating auxin redistribution via alterations in PIN1 degradation and PIN2 biosynthesis.
RESUMO
Bacterial wilt, caused by Ralstonia solanacearum, one of the most destructive phytopathogens, leads to significant annual crop yield losses. Type III effectors (T3Es) mainly contribute to the virulence of R. solanacearum, usually by targeting immune-related proteins. Here, we clarified the effect of a novel E3 ubiquitin ligase (NEL) T3E, RipAW, from R. solanacearum on pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) and further explored its action mechanism. In the susceptible host Arabidopsis thaliana, we monitored the expression of PTI marker genes, flg22-induced ROS burst, and callose deposition in RipAW- and RipAWC177A-transgenic plants. Our results demonstrated that RipAW suppressed host PTI in an NEL-dependent manner. By Split-Luciferase Complementation, Bimolecular Fluorescent Complimentary, and Co-Immunoprecipitation assays, we further showed that RipAW associated with three crucial components of the immune receptor complex, namely FLS2, XLG2, and BIK1. Furthermore, RipAW elevated the ubiquitination levels of FLS2, XLG2, and BIK1, accelerating their degradation via the 26S proteasome pathway. Additionally, co-expression of FLS2, XLG2, or BIK1 with RipAW partially but significantly restored the RipAW-suppressed ROS burst, confirming the involvement of the immune receptor complex in RipAW-regulated PTI. Overall, our results indicate that RipAW impairs host PTI by disrupting the immune receptor complex. Our findings provide new insights into the virulence mechanism of R. solanacearum.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Ralstonia solanacearum , Complexo Antígeno-Anticorpo , Reconhecimento da Imunidade Inata , Espécies Reativas de Oxigênio , Imunoprecipitação , Receptores Imunológicos , Proteínas Serina-Treonina Quinases , Proteínas de Arabidopsis/genéticaRESUMO
RING-finger-type ubiquitin E3 ligase Constitutively Photomorphogenic 1 (COP1) and floral integrators such as FLOWERING LOCUS T (FT), TWIN SISTER OF FT (TSF) and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS1 (SOC1) have been identified as regulators of stomatal movement. However, little is known about their roles and relationship in dark-induced stomatal closure. Here, we demonstrated that COP1 is required for dark-induced stomatal closure using cop1 mutant. The cop1 mutant closed stomata in response to exogenous nitric oxide (NO) but not hydrogen peroxide (H2O2), and H2O2 but not NO accumulated in cop1 in darkness, further indicating that COP1 acts downstream of H2O2 and upstream of NO in dark-induced stomatal closure. Expression of FT, TSF and SOC1 in wild-type (WT) plants decreased significantly with dark duration time, but this process was blocked in cop1. Furthermore, ft, tsf, and soc1 mutants accumulated NO and closed stomata faster than WT plants in response to darkness. Altogether, our results indicate that COP1 transduces H2O2 signaling, promotes NO accumulation in guard cells by suppressing FT, TSF and SOC1 expression, and consequently leads to stomatal closure in darkness. These findings add new insights into the mechanisms of dark-induced stomatal closure.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Óxido Nítrico/metabolismo , Estômatos de Plantas/metabolismo , Peróxido de Hidrogênio/farmacologia , Peróxido de Hidrogênio/metabolismo , Regulação da Expressão Gênica de Plantas , Proteína de Ligação a Fosfatidiletanolamina/genéticaRESUMO
Nitrate is the preferred nitrogen source for plants and plays an important role in plant growth and development. Under various soil stresses, plants reallocate nitrate to roots to promote stress tolerance through the ethylene-ethylene response factors (ERFs)-nitrate transporter (NRT) signaling module. As a light signal, ultraviolet B (UV-B) also stimulates the production of ethylene. However, whether UV-B regulates nitrate reallocation in plants via ethylene remains unknown. Here, we found that UV-B-induced expression of ERF1B, ORA59, ERF104, and NRT1.8 in both Arabidopsis shoots and roots as well as nitrate reallocation from hypocotyls to leaves and roots were impaired in ethylene signaling mutants for Ethylene Insensitive2 (EIN2) and EIN3. UV-B-induced NRT1.8 expression and nitrate reallocation to leaves and roots were also inhibited in the triple mutants for ERF1B, ORA59, and ERF104. Deletion of NRT1.8 impaired UV-B-induced nitrate reallocation to both leaves and roots. Furthermore, UV-B promoted ethylene release in both shoots and roots by enhancing the gene expression and enzymatic activities of ethylene biosynthetic enzymes only in shoots. These results show that ethylene acts as a local and systemic signal to mediate UV-B-induced nitrate reallocation from Arabidopsis hypocotyls to both leaves and roots via regulating the gene expression of the ERFs-NRT1.8 signaling module.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Proteínas de Transporte de Ânions/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Etilenos/metabolismo , Fator VIII/genética , Regulação da Expressão Gênica de Plantas , Mutação , Nitratos/metabolismo , Óxidos de Nitrogênio/metabolismo , Folhas de Planta/genética , Folhas de Planta/metabolismo , Proteínas de Plantas/genética , Raízes de Plantas/metabolismoRESUMO
Heterotrimeric G proteins function as key players in guard cell signaling to many stimuli, including ultraviolet B (UV-B) and ethylene, but whether guard cell G protein signaling is activated by the only one potential G protein-coupled receptor, GCR1, is still unclear. Here, we found that gcr1 null mutants showed defects in UV-B- and ethylene-induced stomatal closure and production of reactive oxygen species (ROS) and nitric oxide (NO) in guard cells, but these defects could be rescued by the application of a Gα activator or overexpression of a constitutively active form of Gα subunit GPA1 (cGPA1). Moreover, the exogenous application of hydrogen peroxide (H2O2) or NO triggered stomatal closure in gcr1 mutants and cGPA1 transgenic plants in the absence or presence of UV-B or ethylene, but exogenous ethylene could not rescue the defect of gcr1 mutants in UV-B-induced stomatal closure, and gcr1 mutants did not affect UV-B-induced ethylene production in Arabidopsis leaves. These results indicate that GCR1 positively controls UV-B- and ethylene-induced stomatal closure by activating GPA1-dependent ROS and NO production in guard cells and that ethylene acts upstream of GCR1 to transduce UV-B guard cell signaling, which establishes the existence of a classic paradigm of G protein signaling in guard cell signaling to UV-B and ethylene.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Etilenos/metabolismo , Etilenos/farmacologia , Subunidades alfa de Proteínas de Ligação ao GTP/metabolismo , Peróxido de Hidrogênio/metabolismo , Peróxido de Hidrogênio/farmacologia , Óxido Nítrico/metabolismo , Estômatos de Plantas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptores Acoplados a Proteínas G/metabolismoRESUMO
Mitogen-activated protein kinases (MPKs) play essential roles in guard cell signaling, but whether MPK cascades participate in guard cell ethylene signaling and interact with hydrogen peroxide (H2 O2 ), nitric oxide (NO), and ethylene-signaling components remain unclear. Here, we report that ethylene activated MPK3 and MPK6 in the leaves of wild-type Arabidopsis thaliana as well as ethylene insensitive2 (ein2), ein3, nitrate reductase1 (nia1), and nia2 mutants, but this effect was impaired in ethylene response1 (etr1), nicotinamide adenine dinucleotide phosphate oxidase AtrbohF, mpk kinase1 (mkk1), and mkk3 mutants. By contrast, the constitutive triple response1 (ctr1) mutant had constitutively active MPK3 and MPK6. Yeast two-hybrid, bimolecular fluorescence complementation, and pull-down assays indicated that MPK3 and MPK6 physically interacted with MKK1, MKK3, and the C-terminal region of EIN2 (EIN2 CEND). mkk1, mkk3, mpk3, and mpk6 mutants had typical levels of ethylene-induced H2 O2 generation but impaired ethylene-induced EIN2 CEND cleavage and nuclear translocation, EIN3 protein accumulation, NO production in guard cells, and stomatal closure. These results show that the MKK1/3-MPK3/6 cascade mediates ethylene-induced stomatal closure by functioning downstream of ETR1, CTR1, and H2 O2 to interact with EIN2, thereby promoting EIN3 accumulation and EIN3-dependent NO production in guard cells.
Assuntos
Proteínas de Arabidopsis/metabolismo , Proteínas de Ligação a DNA/metabolismo , Etilenos/farmacologia , MAP Quinase Quinase 1/metabolismo , MAP Quinase Quinase 3/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Estômatos de Plantas/efeitos dos fármacos , Estômatos de Plantas/metabolismo , Receptores de Superfície Celular/metabolismo , Fatores de Transcrição/metabolismo , Arabidopsis/efeitos dos fármacos , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Ligação a DNA/genética , MAP Quinase Quinase 1/genética , MAP Quinase Quinase 3/genética , Proteínas Quinases Ativadas por Mitógeno/genética , Receptores de Superfície Celular/genética , Fatores de Transcrição/genéticaRESUMO
OBJECTIVE: To investigate the current status of antibiotic use for very and extremely low birth weight (VLBW/ELBW) infants in neonatal intensive care units (NICUs) of Hunan Province. METHODS: The use of antibiotics was investigated in multiple level 3 NICUs of Hunan Province for VLBW and ELBW infants born between January, 2017 and December, 2017. RESULTS: The clinical data of 1â442 VLBW/ELBW infants were collected from 24 NICUs in 2017. The median antibiotic use duration was 17 days (range: 0-86 days), accounting for 53.0% of the total length of hospital stay. The highest duration of antibiotic use was up to 91.4% of the total length of hospital stay, with the lowest at 14.6%. In 16 out of 24 NICUs, the antibiotic use duration was accounted for more than 50.0% of the hospitalization days. There were 113 cases with positive bacterial culture grown in blood or cerebrospinal fluid, making the positive rate of overall bacterial culture as 7.84%. The positive rate of bacterial culture in different NICUs was significantly different from 0% to 14.9%. The common isolated bacterial pathogens Klebsiella pneumoniae was 29 cases (25.7%); Escherichia coli 12 cases (10.6%); Staphylococcus aureus 3 cases (2.7%). The most commonly used antibiotics were third-generation of cephalosporins, accounting for 41.00% of the total antibiotics, followed by penicillins, accounting for 32.10%, and followed by carbapenems, accounting for 13.15%. The proportion of antibiotic use time was negatively correlated with birth weight Z-score and the change in weight Z-score between birth and hospital discharge (rs=-0.095, -0.151 respectively, P<0.01), positively correlated with death/withdrawal of care (rs=0.196, P<0.01). CONCLUSIONS: Antibiotics used for VLBW/ELBW infants in NICUs of Hunan Province are obviously prolonged in many NICUs. The proportion of routine use of third-generation of cephalosporins and carbapenems antibiotics is high among the NICUs.
Assuntos
Recém-Nascido de Peso Extremamente Baixo ao Nascer , Antibacterianos , Peso ao Nascer , Humanos , Lactente , Recém-Nascido , Unidades de Terapia Intensiva Neonatal , Inquéritos e QuestionáriosRESUMO
Neuropathic pain is a kind of pain caused by primary or secondary impairment or dysfunction of peripheral or central nervous system. Patients with neuropathic pain were often with poor clinical outcome. We screened the differentially expressed genes between sciatic nerve injury and dorsal root ganglion gene in the sham operation model. Microarray and the spared nerve injury module were used to explore the molecular mechanism of neuropathic pain by injuries and the differentially expressed genes (DEGs) were identified out. Besides, the bioinformatics methods were used to figure out the signaling pathways and expression regulation pattern these DEGs were enriched in, which may provide a basis for the molecular research and medicine target of therapy. Besides, protein-protein interaction network analysis was performed on these selected intersection genes. A total of 40 DEGs were screened out and only pctp gene was down-regulated, the left 39 genes were all up-regulated. Then, GO and KEGG enrichment analysis were performed on these intersection genes by DAVID software. Furthermore, protein-protein interaction network analysis was used to analyze the critical genes of neuropathic pain. Finally, four genes, that is, Jun, Gal, Cd74, and C1qb were identified to have strong interactions with other genes, which may function as the prognostic and predictive genes of neuropathic pain caused by peripheral injuries. Our results suggested that four differentially expressed genes, Jun, Gal, Cd74, and C1qb, had the potential to serve as prognostic or predictive markers for neuropathic pain, suggesting a potential application in the improvement of prognostic tools and treatments.
Assuntos
Antígenos de Diferenciação de Linfócitos B/metabolismo , Proteínas de Transporte/metabolismo , Galanina/metabolismo , Antígenos de Histocompatibilidade Classe II/metabolismo , Proteínas Mitocondriais/metabolismo , Neuralgia/metabolismo , Traumatismos dos Nervos Periféricos/metabolismo , Proteínas Proto-Oncogênicas c-jun/metabolismo , Biomarcadores/metabolismo , Regulação da Expressão Gênica , Humanos , Neuralgia/patologia , SoftwareRESUMO
Ultraviolet B (UV-B) radiation induces the activation of MITOGEN-ACTIVATED PROTEIN KINASE PHOSPHATASE1 (MKP1) and its targets MPK3 and MPK6, but whether they participate in UV-B guard cell signaling is not clear. Here, evidence shows that UV-B-induced stomatal closure in Arabidopsis (Arabidopsis thaliana) is antagonistically regulated by MKP1 and MPK6 via modulating hydrogen peroxide (H2O2)-induced nitric oxide (NO) production in guard cells. The mkp1 mutant was hypersensitive to UV-B-induced stomatal closure and NO production in guard cells but not to UV-B-induced H2O2 production, suggesting that MKP1 negatively regulates UV-B-induced stomatal closure via inhibiting NO generation in guard cells. Moreover, MPK3 and MPK6 were activated by UV-B in leaves of the wild type and hyperactivated in the mkp1 mutant, but the UV-B-induced activation of MPK3 and MPK6 was largely inhibited in mutants for ATRBOHD and ATRBOHF but not in mutants for NIA1 and NIA2 mpk6 mutants showed defects of UV-B-induced NO production and stomatal closure but were normal in UV-B-induced H2O2 production, while stomata of mpk3 mutants responded to UV-B just like those of the wild type. The defect of UV-B-induced stomatal closure in mpk6 mutants was rescued by exogenous NO but not by exogenous H2O2 Furthermore, double mutant mkp1/mpk6 and the single mutant mpk6 showed the same responses to UV-B in terms of either stomatal movement or H2O2 and NO production. These data indicate that MPK6, but not MPK3, positively regulates UV-B-induced stomatal closure via acting downstream of H2O2 and upstream of NO, while MKP1 functions negatively in UV-B guard cell signaling via down-regulation of MPK6.
Assuntos
Proteínas de Arabidopsis/metabolismo , Óxido Nítrico/metabolismo , Estômatos de Plantas/fisiologia , Arabidopsis/citologia , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , Peróxido de Hidrogênio/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Fosfatases da Proteína Quinase Ativada por Mitógeno/genética , Fosfatases da Proteína Quinase Ativada por Mitógeno/metabolismo , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Mutação , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Células Vegetais/metabolismo , Células Vegetais/efeitos da radiação , Estômatos de Plantas/efeitos da radiação , Proteínas Tirosina Fosfatases/genética , Proteínas Tirosina Fosfatases/metabolismo , Transdução de Sinais , Raios UltravioletaRESUMO
Heterotrimeric G proteins function as key players in hydrogen peroxide (H2O2) production in plant cells, but whether G proteins mediate ethylene-induced H2O2 production and stomatal closure are not clear. Here, evidences are provided to show the Gα subunit GPA1 as a missing link between ethylene and H2O2 in guard cell ethylene signalling. In wild-type leaves, ethylene-triggered H2O2 synthesis and stomatal closure were dependent on activation of Gα. GPA1 mutants showed the defect of ethylene-induced H2O2 production and stomatal closure, whereas wGα and cGα overexpression lines showed faster stomatal closure and H2O2 production in response to ethylene. Ethylene-triggered H2O2 generation and stomatal closure were impaired in RAN1, ETR1, ERS1 and EIN4 mutants but not impaired in ETR2 and ERS2 mutants. Gα activator and H2O2 rescued the defect of RAN1 and EIN4 mutants or etr1-3 in ethylene-induced H2O2 production and stomatal closure, but only rescued the defect of ERS1 mutants or etr1-1 and etr1-9 in ethylene-induced H2O2 production. Stomata of CTR1 mutants showed constitutive H2O2 production and stomatal closure, but which could be abolished by Gα inhibitor. Stomata of EIN2, EIN3 and ARR2 mutants did not close in responses to ethylene, Gα activator or H2O2, but do generate H2O2 following challenge of ethylene or Gα activator. The data indicate that Gα mediates ethylene-induced stomatal closure via H2O2 production, and acts downstream of RAN1, ETR1, ERS1, EIN4 and CTR1 and upstream of EIN2, EIN3 and ARR2. The data also show that ETR1 and ERS1 mediate both ethylene and H2O2 signalling in guard cells.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Etilenos/farmacologia , Peróxido de Hidrogênio/metabolismo , Reguladores de Crescimento de Plantas/farmacologia , Transdução de Sinais , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Subunidades alfa de Proteínas de Ligação ao GTP/genética , Subunidades alfa de Proteínas de Ligação ao GTP/metabolismo , Regulação da Expressão Gênica de Plantas , Mutação , Folhas de Planta/genética , Estômatos de Plantas/genética , Estômatos de Plantas/fisiologia , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismoRESUMO
Heterotrimeric G proteins have been shown to transmit ultraviolet B (UV-B) signals in mammalian cells, but whether they also transmit UV-B signals in plant cells is not clear. In this paper, we report that 0.5 W m(-2) UV-B induces stomatal closure in Arabidopsis (Arabidopsis thaliana) by eliciting a cascade of intracellular signaling events including Gα protein, hydrogen peroxide (H2O2), and nitric oxide (NO). UV-B triggered a significant increase in H2O2 or NO levels associated with stomatal closure in the wild type, but these effects were abolished in the single and double mutants of AtrbohD and AtrbohF or in the Nia1 mutants, respectively. Furthermore, we found that UV-B-mediated H2O2 and NO generation are regulated by GPA1, the Gα-subunit of heterotrimeric G proteins. UV-B-dependent H2O2 and NO accumulation were nullified in gpa1 knockout mutants but enhanced by overexpression of a constitutively active form of GPA1 (cGα). In addition, exogenously applied H2O2 or NO rescued the defect in UV-B-mediated stomatal closure in gpa1 mutants, whereas cGα AtrbohD/AtrbohF and cGα nia1 constructs exhibited a similar response to AtrbohD/AtrbohF and Nia1, respectively. Finally, we demonstrated that Gα activation of NO production depends on H2O2. The mutants of AtrbohD and AtrbohF had impaired NO generation in response to UV-B, but UV-B-induced H2O2 accumulation was not impaired in Nia1. Moreover, exogenously applied NO rescued the defect in UV-B-mediated stomatal closure in the mutants of AtrbohD and AtrbohF. These findings establish a signaling pathway leading to UV-B-induced stomatal closure that involves GPA1-dependent activation of H2O2 production and subsequent Nia1-dependent NO accumulation.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Subunidades alfa de Proteínas de Ligação ao GTP/metabolismo , Peróxido de Hidrogênio/metabolismo , Óxido Nítrico/metabolismo , Estômatos de Plantas/fisiologia , Estômatos de Plantas/efeitos da radiação , Raios Ultravioleta , Arabidopsis/enzimologia , Arabidopsis/efeitos da radiação , Peróxido de Hidrogênio/farmacologia , NADPH Oxidases/metabolismo , Óxido Nítrico/biossíntese , Estômatos de Plantas/citologia , Estômatos de Plantas/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacosRESUMO
To alleviate toxicological effect induced by cadmium in mungbean seedlings, seeds were divided into four groups: The controls groups (CK, without treatment), magnetic field treated groups (MF), cadmium treated groups (CS), and magnetic field treated followed by cadmium treated groups (MF + CS).The results showed: (i) Compared with the controls, cadmium stress resulted in enhancing in the concentration of malondialdehyde, H(2)O(2) and O(2-), and the conductivity of electrolyte leakage while decreasing in the nitrice oxide synthase (NOS) activity, the concentration of nitrice oxide (NO), chlorophyll and total carbon and nitrogen, the net photosynthetic rate, the stomatal conductance, the transpiration rate, the water use efficiency, the lateral number and seedlings growth except for intercellular CO(2) concentration increase. However, the seedlings treated with 600 mT magnetic field followed by cadmium stress the concentration of malondialdehyde, H(2)O(2) and O(2-), and the conductivity of electrolyte leakage decreased, while the above mentioned NO concentration, NOS activity, photosynthesis and growth parameters increased compared to cadmium stress alone. (ii) Compared with the cadmium stress (CS), the seedling growth were inhibited when the seeds were treated with NO scavenger (hemoglobin, HB) and inhibition of NO generating enzyme (sodium tungstate, ST), conversely, the seedling growth were improved by the NO donor sodium nitroprusside (SNP) and CaCl(2). In the case of the HB and ST treatment followed by magnetic field and then the seedling subjected to CS, the seedlings growth was better than that of hemoglobin (HB) followed by CS and ST followed by CS. The seeds were treated with SNP and CaCl(2) followed by MF, and then subjected to CS, the seedlings growth were better than that of SNP followed by CS, and CaCl(2) followed by CS. These results suggested that magnetic field compensates for the toxicological effects of cadmium exposure are related to NO signal.
Assuntos
Cádmio/toxicidade , Fabaceae/efeitos dos fármacos , Magnetismo , Poluentes do Solo/toxicidade , Fabaceae/metabolismo , Fabaceae/fisiologia , Peróxido de Hidrogênio/metabolismo , Malondialdeído/metabolismoRESUMO
Although the UV RESISTANCE LOCUS 8 (UVR8)-CONSTITUTIVELY PHOTOMORPHOGENIC1 (COP1)-ELONGATED HYPOCOTYL5 (HY5) signaling pathway, ethylene, hydrogen peroxide (H2O2), and nitric oxide (NO) all participate in ultraviolet-B (UV-B)-triggered stomatal closing, their interrelationship is not clear. Here, we found that UV-B-induced the expression of ethylene biosynthetic genes, production of ethylene, H2O2, and NO, and stomata closing were impaired in uvr8, cop1, and hy5 mutants. UV-B-induced NO production and stomata closing were also defective in mutants for ETHYLENE RESPONSE 1 (ETR1), ETHYLENE INSENSITIVE 2 (EIN2), and EIN3, but UV-B-triggered H2O2 generation was only inhibited in etr1. In either the absence or presence of UV-B, ethylene triggered H2O2 production but not NO generation and stomatal closure in cop1 and hy5, and stomata closing in cop1 and hy5 was induced by NO but not H2O2. Moreover, NO production and stomatal closure were constitutively caused by over-expression of COP1 or HY5 in ein2 and ein3, but not by over-expression of EIN2 or EIN3 in cop1 and hy5. Our data indicate that the UVR8-COP1-HY5 signaling module mediates UV-B-induced ethylene production, ethylene is then perceived by ETR1 to induce H2O2 synthesis. H2O2 induces NO generation and subsequent stomata closing via an EIN2, EIN3, COP1, and HY5-dependent pathway(s).
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Proteínas Cromossômicas não Histona/metabolismo , Etilenos/metabolismo , Peróxido de Hidrogênio/metabolismo , Óxido Nítrico/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Transdução de Sinais , Arabidopsis/fisiologia , Arabidopsis/efeitos da radiação , Proteínas de Arabidopsis/genética , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Proteínas Cromossômicas não Histona/genética , Expressão Gênica , Mutação , Estômatos de Plantas/genética , Estômatos de Plantas/fisiologia , Estômatos de Plantas/efeitos da radiação , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Raios UltravioletaRESUMO
Stomata are a key land plant innovation that permit the regulation of gaseous exchanges between the plant interior and the surrounding environment. By opening or closing, stomata regulate transpiration of water though the plant; and these actions are coordinated with acquisition of CO2 for photosynthesis. Stomatal movement is controlled by various environmental and physiological factors and associates with multiple intracellular activities, among which the dynamic remodeling of vacuoles plays a crucial role. Phosphatidylinositol 3,5-bisphosphate [PI(3,5)P2] is critical for dynamic remodeling of vacuoles. Its production requires a PI(3,5)P2-metabolizing complex consisting of FAB1/PIKfyve kinases, SAC phosphatases, and the scaffolding protein VAC14. Although genetic or pharmacological downregulation of PI(3,5)P2 causes hyposensitivity to ABA-induced stomatal closure, whether the effect of PI(3,5)P2 on stomatal movement is cell-autonomous and the physiological consequences of its reduction were unclear. We report that downregulating Arabidopsis VAC14 specifically in guard cells by artificial microRNAs (amiR-VAC14) results in enlarged guard cells and hyposensitivity to ABA- and dark-induced stomatal closure. Vacuolar fission during stomatal closure is compromised by downregulating VAC14 in guard cells. Exogenous application of PI(3,5)P2 rescued the amiR-VAC14 phenotype whereas PI(3,5)P2 inhibitor YM201636 caused wild-type plants to have inhibited stomatal closure. We further show that downregulating VAC14 specifically in guard cells impairs drought tolerance, suggestive of a key role of guard cell-produced PI(3,5)P2 in plant fitness.
RESUMO
Both salicylic acid (SA) and ethylene induce stomatal closure and positively regulate stomatal immunity, but their interactions in guard cell signaling are unclear. Here, we observed that SA induced the expression of ethylene biosynthetic genes; the production of ethylene, reactive oxygen species (ROS) and nitric oxide (NO); and stomatal closure in Arabidopsis thaliana. However, SA-induced stomatal closure was inhibited by an ethylene biosynthetic inhibitor and mutations in ethylene biosynthetic genes, ethylene-signaling genes [RESPONSE TO ANTAGONIST 1 (RAN1), ETHYLENE RESPONSE 1 (ETR1), ETHYLENE INSENSITIVE 2 (EIN2), EIN3 and ARABIDOPSIS RESPONSE REGULATOR 2 (ARR2)], NADPH oxidase genes [ATRBOHD and ATRBOHF], and nitrate reductase genes (NIA1 and NIA2). Furthermore, SA-triggered ROS production in guard cells was impaired in ran1, etr1, AtrbohD and AtrbohF, but not in ein2, ein3 or arr2. SA-triggered NO production was impaired in all ethylene-signaling mutants tested and in nia1 and nia2. The stomata of mutants for CONSTITUTIVE TRIPLE RESPONSE1 (CTR1) showed constitutive ROS and NO production and closure. These results indicate that ethylene mediates SA-induced stomatal closure by activating ATRBOHD/F-mediated ROS synthesis in an RAN1-, ETR1- and CTR1-dependent manner. This in turn induces NIA1/2-mediated NO production and subsequent stomatal closure via the ETR1, EIN2, EIN3 and ARR2-dependent pathway(s).
Assuntos
Arabidopsis/metabolismo , Óxido Nítrico/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas de Arabidopsis/metabolismo , Etilenos/metabolismo , NADPH Oxidases/metabolismo , Estômatos de Plantas/metabolismo , Ácido Salicílico/metabolismoRESUMO
The role of nitric oxide (NO) in the ultraviolet-B radiation (UV-B)-induced reduction of in vitro pollen germination and tube growth of Paulownia tomentosa Steud. was studied. Results showed that exposure of the pollen to 0.4 and 0.8 W m(-2) UV-B radiation for 2 h resulted in not only the reduction of pollen germination and tube growth but also the enhancement of NO synthase (NOS, EC 1.14.13.39) activity and NO production in pollen grain and tube. Also, exogenous NO donors sodium nitroprusside and S-nitrosoglutathione inhibited both pollen germination and tube growth in a dose-dependence manner. NOS inhibitor N(G)-nitro-l-Arg-methyl eater (l-NAME) and NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO) not only largely prevented the NO generation but also partly reversed the UV-B-inhibited pollen germination and tube growth. These results indicate that UV-B radiation inhibits pollen germination and tube growth partly via promoting NO production in pollen grain and tube by a NOS-like enzyme. Additionally, a guanylyl cyclase inhibitor 6-anilino-5,8-quinolinequinone (LY-83583) prevented both the UV-B- and NO donors-inhibited pollen germination and tube growth, suggesting that the NO function is mediated by cyclic guanosine 5'-monophosphate. However, the effects of c-PTIO, l-NAME and LY-83583 on the UV-B-inhibited pollen germination and tube growth were only partial, suggesting that there are NO-independent pathways in UV-B signal networks.
Assuntos
Óxido Nítrico/metabolismo , Pólen/fisiologia , Pólen/efeitos da radiação , Raios Ultravioleta , GMP Cíclico/metabolismo , Magnoliopsida/efeitos dos fármacos , Magnoliopsida/metabolismo , Magnoliopsida/efeitos da radiação , NG-Nitroarginina Metil Éster/farmacologia , Doadores de Óxido Nítrico/farmacologia , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase/metabolismo , Nitroprussiato/farmacologia , Pólen/efeitos dos fármacos , Resinas Vegetais/metabolismo , S-Nitrosoglutationa/farmacologiaRESUMO
In two mung bean cultivars (Phaseolus raditus L. cv. 'Qindou-20' and 'Zhonglv-1') with different sensitivities to UV-B grown in growth chamber under supplemental or no supplemental UV-B radiation (0.4 W/m(2)) with or without 0.4% NaCl, the effects of NaCl stress on UV-B-induced DNA damage and repair were studied. The results showed that, under NaCl stress, (i) CPD accumulation was lower in the tolerant cultivar 'Zhonglv-1' but was the same in the sensitive cultivar 'Qindou-20', (ii) CPD formation in both cultivars was weakened, (iii) the photorepair and dark repair capacity were higher in the tolerant cultivar and (iv) the photorepair was weakened and dark repair capacity did not change in the sensitive cultivar. There was a negative correlation between susceptibility of CPD formation and levels of UV-absorbing compounds. These results demonstrate that NaCl stress can affect not only the susceptibility to CPD formation, but also the capacities for photorepair and dark repair of DNA, which together result in the change in UV-B-induced CPD accumulation and thereby that in sensitivity of plant to UV-B. The results also suggest that the differences in susceptibilities to CPD formation are due to the differences in levels of UV-absorbing compounds.
Assuntos
Fabaceae/efeitos dos fármacos , Fabaceae/metabolismo , Dímeros de Pirimidina/metabolismo , Dímeros de Pirimidina/efeitos da radiação , Cloreto de Sódio/farmacologia , Raios Ultravioleta , Ensaio de Imunoadsorção Enzimática , Fabaceae/efeitos da radiaçãoRESUMO
Pharmacological data have suggested the involvement of mitogen-activated protein kinase (MPK) cascades in dark-induced stomatal closure, but which specific MPK cascade participates in the darkness guard cell signaling and its relationship with hydrogen peroxide (H2O2) and nitric oxide (NO) remain unclear. In this paper, we observed that darkness induced activation of MPK6 in leaves of wild-type Arabidopsis (Arabidopsis thaliana) and mutants for nitrate reductase 1 (NIA1), but this effect was inhibited in mutants for MPK Kinase 1 (MEK1) and ATRBOHD/F. Mutants for MEK1, MPK6 and NIA1 showed defect of dark-induced NO production in guard cells and stomatal closure, but were normal in the dark-induced H2O2 generation, while stomata of mutant AtrbohD/F showed defect of dark-induced H2O2 and NO production and subsequent closure. Moreover, exogenous NO rescued the defect of dark-induced stomatal closure in mutants of AtrbohD/F, mek1 and mpk6, while exogenous H2O2 could not rescue the defect of dark-induced stomatal closure in mutants of mek1, mpk6 and nia1. These genetic and biochemical evidences not only show that MEK1-MPK6 cascade, AtRBOHD/F-dependent H2O2 and NIA1-dependent NO are all involved in dark-induced stomatal closure in Arabidopsis, also indicate that MEK1-MPK6 cascade functions via working downstream of H2O2 and upstream of NO.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Arabidopsis/fisiologia , Escuridão , Peróxido de Hidrogênio/metabolismo , MAP Quinase Quinase 1/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Óxido Nítrico/metabolismo , Estômatos de Plantas/metabolismo , Estômatos de Plantas/fisiologia , Proteínas de Arabidopsis/genética , MAP Quinase Quinase 1/genética , Proteínas Quinases Ativadas por Mitógeno/genética , Nitrato Redutase/genética , Nitrato Redutase/metabolismoRESUMO
Stomatal and nonstomatal limitations of photosynthesis in mung bean (Phaseolus radiatus L.) leaves under the combination of 0.35 W/m(2) UV-B radiation and 0.4% NaCl stress were studied. Separated or combined treatments of enhanced UV-B radiation and NaCl stress all resulted in a decrease in net photosynthetic rate, stomatal conductance, photosynthetic ability, efficiency of CO(2) carboxylation and Rubisco content, and the extent of those decreases enhanced obviously with the increase of treatment days. Intercellular CO(2)concentration tended to decrease in short period, followed by a gradual increase and to be higher than the control on the seventh day after treatment under NaCl stress alone, while a tendency of gradual increase was observed under enhanced UV-B radiation alone or two stresses combined and the concentration was higher than the control after three day of treatments. By contrast, the tendency of stomatal limitation value was just reverse to the intercellular CO(2) concentration, but the stomatal limitation value was always higher than control under all stress conditions (did not include that on the fifth day under the combined stresses). As compared with an individual stress the effect of combined stress on the above parameters was more serious. These results indicate that under all stress conditions the inhibition of photosynthesis in mung bean leaves is the results of both stomatal and nonstomatal limitations. And the stomatal limitation is dominant in short period, nonstomatal limitation becomes the dominant one in longer period. The decrease in Rubisco content leads to nonstomatal limitation of photosynthesis under all stresses.
Assuntos
Phaseolus/metabolismo , Fotossíntese , Folhas de Planta/metabolismo , Cloreto de Sódio/farmacologia , Phaseolus/efeitos dos fármacos , Phaseolus/efeitos da radiação , Ribulose-Bifosfato Carboxilase/análise , Raios UltravioletaRESUMO
To understand the role of H(2)O(2) in the damaging effect of UV-B radiation on ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) content, the time courses of changes in Rubisco and H(2)O(2) contents under UV-B treatment were measured in the primary leaves of two mung bean cultivars (Phaseolus raditus L. cv. Zhonglv-1 and Qindou-20). A 0.4 W/m(2) UV-B treatment caused a drop of the former to about 1/3 and a 30%-40% rise in H(2)O(2) content in a week in both cultivars but with Qindou-20 showing stronger responses. Such higher rate of increase in proteolytic activity under UV-B treatment implies that UV-B lowers Rubisco by activating proteolytic enzymes which hydrolizes Rubisco. Scavenging of H(2)O(2) by exogenously added ascorbic acid (AsA) significantly inhibited the UV-B-induced decrease in Rubisco amount and increases in H(2)O(2) level and proteolytic activity. It is shown that H(2)O(2) participates in the UV-B-induced reduction of Rubisco amount. It is suggested that H(2)O(2) stimulates the degradation of Rubisco by activation of proteolytic systems.