Polyamines （PA）, polyamine oxidases, copper amine oxidases, and nitric oxide （NO） play important roles in physiology and stress responses in plants. NO biosynthesis as a result of catabolism of PA by polyamine oxi...Polyamines （PA）, polyamine oxidases, copper amine oxidases, and nitric oxide （NO） play important roles in physiology and stress responses in plants. NO biosynthesis as a result of catabolism of PA by polyamine oxidases and copper amine oxidases may explain in part PA-mediated responses. Involvement of a copper amine oxidase gene, COPPER AMINE OXIDASEI （CuAO1）, of Arabidopsis was tested for its role in stress responses using the knockouts cuao1-1 and cuaol-2. PA-induced and ABA-induced NO production investigated bY fluorometry and fluorescence microscopy showed that the cuaol-1 and cuaol-2 are impaired in NO production, suggesting a function of CuAO1 in PA and ABA-mediated NO production. Furthermore, we found a PA-dependent increase in protein S-nitrosylation. The addition of PA and ABA also resulted in H2O2 increases, cuao1-1 and cuao1-2 showed less sensitivity to exogenous ABA supplementation during germination, seedling establishment, and root growth inhibition as compared to wild-type. In response to ABA treatment, expression levels of the stress-responsive genes RD29A and ADH1 were significantly lower in the knockouts. These observations characterize cuao1-1 and cuao1-2 as ABA-insensitive mutants. Taken together, our findings extend the ABA signal transduction network to include CuAO1 as one potential contributor to enhanced NO production by ABA.展开更多
The polar, sub-cellular localization of PIN auxin efflux carriers determines the direction of intercellular auxin flow, thus defining the spatial aspect of auxin signalling. Dynamic, transcytosis-like relocalizations ...The polar, sub-cellular localization of PIN auxin efflux carriers determines the direction of intercellular auxin flow, thus defining the spatial aspect of auxin signalling. Dynamic, transcytosis-like relocalizations of PIN proteins occur in response to external and internal signals, integrating these signals into changes in auxin distribution. Here, we examine the cellular and molecular mechanisms of polar PIN delivery and transcytosis. The mechanisms of the ARF-GEF-dependent polar targeting and transcytosis are well conserved and show little variations among diverse Arabidopsis ecotypes consistent with their fundamental importance in regulating plant development. At the cellular level, we refine previous findings on the role of the actin cytoskeleton in apical and basal PIN targeting, and identify a previously unknown role for microtubules, specifically in basal targeting. PIN protein delivery to different sides of the cell is mediated by ARFdependent trafficking with a previously unknown complex level of distinct ARF-GEF vesicle trafficking regulators. Our data suggest that alternative recruitment of PIN proteins by these distinct pathways can account for cell type- and cargo-specific aspects of polar targeting, as well as for polarity changes in response to different signals. The resulting dynamic PIN positioning to different sides of cells defines a three-dimensional pattern of auxin fluxes within plant tissues.展开更多
As the first and rate-limiting enzyme of proline degradation, PROLINE DEHYDROGENASE1 （PDH1） is tightly regulated during plant stress responses, including induction under hypoosmolarity and repression under water def...As the first and rate-limiting enzyme of proline degradation, PROLINE DEHYDROGENASE1 （PDH1） is tightly regulated during plant stress responses, including induction under hypoosmolarity and repression under water deficit. The plant receptor histidine kinases AHKs, elements of the two-component system （TCS） in Arabidopsis thaliana, are pro- posed to function in water stress responses by regulating different stress-responsive genes. However, little information is available concerning AHK phosphorelay-mediated downstream signaling. Here we show that the Arabidopsis type-B response regulator 18 （ARR18） functions as a positive osmotic stress response regulator in Arabidopsis seeds and affects the activity of the PDH1 promoter, known to be controlled by C-group bZlP transcription factors. Moreover, direct physical interaction of ARR18 with bZIP63 was identified and shown to be dependent on phosphorylation of the conserved aspar- tate residue in the ARR18 receiver domain. We further show that bZlP63 itself functions as a negative regulator of seed germination upon osmotic stress. Using reporter gene assays in protoplasts, we demonstrated that ARR18 interaction negatively interferes with the transcriptional activity of bZIP63 on the PDH1 promoter. Our findings provide new insight into the function of ARR18 and bZIP63 as antagonistic regulators of gene expression in Arabidopsis.展开更多
High-temperature stress, like any abiotic stress, impairs the physiology and development of plants, including the stages of seed setting and ripening. We used the Affymetrix 22K Barley1 GeneChip microarray to investig...High-temperature stress, like any abiotic stress, impairs the physiology and development of plants, including the stages of seed setting and ripening. We used the Affymetrix 22K Barley1 GeneChip microarray to investigate the response of developing barley （Hordeum vulgare） seeds, termed caryopses, after 0.5, 3, and 6 h of heat stress exposure; 958 induced and 1122 repressed genes exhibited spatial and temporal expression patterns that provide a detailed insight into the caryopses＇ early heat stress responses. Down-regulation of genes related to storage compound biosynthesis and cell growth provides evidence for a rapid impairment of the caryopsis＇ development. Increased levels of sugars and amino acids were indicative for both production of compatible solutes and feedback-induced accumulation of substrates for storage compound biosynthesis. Metadata analysis identified embryo and endosperm as primary locations of heat stress responses, indicating a strong impact of short-term heat stress on central developmental functions of the caryopsis. A comparison with heat stress responses in Arabidopsis shoots and drought stress responses in barley caryopses identified both conserved and presumably heat- and caryopsis-specific stress-responsive genes. Summarized, our data provide an important basis for further investigation of gene functions in order to aid an improved heat tolerance and reduced losses of yield in barley as a model for cereal crops.展开更多
Besides providing protection against mechanical damage to the root tip, the root cap is involved in the perception and processing of diverse external and internal stimuli resulting in altered growth and development, T...Besides providing protection against mechanical damage to the root tip, the root cap is involved in the perception and processing of diverse external and internal stimuli resulting in altered growth and development, The transduction of these stimuli includes hormonal signaling pathways such as those of auxin, ethylene and cytokinin. Here, we show that the root cap is essential for the ethylene-induced regulation of elongation growth and root hair formation in maize. Exogenously applied ethylene is no longer able to inhibit elongation growth when the root cap has been surgically removed prior to hormone treatment. Reconstitution of the cap positively correlates with the developing capacity of the roots to respond to ethylene again. In contrast, the removal of the root cap does not per se affect growth inhibition controlled by auxin and cytokinin. Furthermore, our semi-quantitative RT-PCR results support earlier findings that the maize root cap is a site of high gene expression activity with respect to sensing and responding to hormones such as ethylene, From these data, we propose a novel function of the root cap which is the establishment of competence to respond to ethylene in the distal zones of the root.展开更多
In order to analyze the signaling function of hydrogen peroxide （H202） production in senescence in more detail, we manipulated intracellular H202 levels in Arabidopsis thaliala （L.） Heynh by using the hydrogen- pe...In order to analyze the signaling function of hydrogen peroxide （H202） production in senescence in more detail, we manipulated intracellular H202 levels in Arabidopsis thaliala （L.） Heynh by using the hydrogen- peroxide-sensitive part of the Escherichia coil transcription regulator OxyR, which was directed to the cytoplasm as well as into the peroxisomes. H202 levels were lowered and senescence was delayed in both transgenic lines, but OxyR was found to be more effective in the cytoplasm. To transfer this knowledge to crop plants, we analyzed oilseed rape plants Brassica napus L. cv. Mozart for H20~ and its scavenging enzymes catalase （CAT） and ascorbate peroxidase （APX） during leaf and plant development. H202 levels were found to increase during bolting and flowering time, but no increase could be observed in the very late stages of senescence. With increasing H202 levels, CAT and APX activities declined, so it is likely that similar mechanisms are used in oilseed rape and Arabidopsis to control H202 levels. Under elevated CO2 conditions, oilseed rape senescence was accelerated and coincided with an earlier increase in H202 levels, indicating that H202 may be one of the signals to inducing senescence in a broader range of Brassicaceae.展开更多
Spectro-microscopy, a combination of fluorescence microscopy with spatially resolved spectroscopic techni- ques, provides new and exciting tools for functional cell biology in living organisms. This review focuses on ...Spectro-microscopy, a combination of fluorescence microscopy with spatially resolved spectroscopic techni- ques, provides new and exciting tools for functional cell biology in living organisms. This review focuses on recent devel- opments in spectro-microscopic applications for the investigation of living plant cells in their native tissue context. The application of spectro-microscopic methods led to the recent discovery of a fast signal response pathway for the brassi- nosteroide receptor BRI1 in the plasma membrane of living plant cells. Moreover, the competence of different plant cell types to respond to environmental or endogenous stimuli was determined in vivo by correlation analysis of different optical and spectroscopic readouts such as fluorescence lifetime （FLT）. Furthermore, a new spectro-microscopic technique, fluorescence intensity decay shape analysis microscopy （FIDSAM）, has been developed. FIDSAM is capable of imaging low- expressed fluorophore-tagged proteins at high spatial resolution and precludes the misinterpretation of autofluorescence artifacts. In addition, FIDSAM provides a very effective and sensitive tool on the basis of F6rster resonance energy transfer （FRET） for the qualitative and quantitative determination of protein-protein interaction. Finally, we report on the quan- titative analysis of the photosystem I and II （PSI/PSII） ratio in the chloroplasts of living Arabidopsis plants at room tem- perature, using high-resolution, spatially resolved fluorescence spectroscopy. With this technique, it was not only possible to measure PSI/PSII ratios, but also to demonstrate the differential competence of wild-type and carbohydrate-deficient plants to adapt the PSI/PSII ratio to different light conditions. In summary, the information content of standard microscopic images is extended by several dimensions by the use of spectro-microscopic approaches. Therefore, novel cell physiolog- ical and molecular topics can be addressed and valuable insights into molecular and subcellula展开更多
Seed size critically affects grain yield of crops and hence represents a key breeding target.The develop-ment of embryo-nourishing endosperm is a key driver of seed expansion.We here report unexpected dual roles of th...Seed size critically affects grain yield of crops and hence represents a key breeding target.The develop-ment of embryo-nourishing endosperm is a key driver of seed expansion.We here report unexpected dual roles of the transcription factor EIN3 in regulating seed size.These EIN3 functions have remained largely undiscovered because they oppose each other.Capitalizing on the analysis of multiple ethylene biosynthesis mutants,we demonstrate that EIN3 represses endosperm and seed development in a pathway regulated by ethylene.We,in addition,provide evidence that EIN3-mediated synergid nucleus disintegration promotes endosperm expansion.Interestingly,synergid nucleus disintegration is not affected in various ethylene biosynthesis mutants,suggesting that this promoting function of EIN3 is inde-pendent of ethylene.Whereas the growth-inhibitory ethylene-dependent EIN3 action appears to be encoded by sporophytic tissue,the growth-promoting role of EIN3 is induced by fertilization,revealing a generation conflict that converges toward the key signaling component EIN3.展开更多
Successful import of hundreds of nucleus-encoded proteins is essential for chloroplast biogenesis. The import of cytosolic precursor proteins relies on the Toc- （translocon at the outer chloroplast membrane） and Ti...Successful import of hundreds of nucleus-encoded proteins is essential for chloroplast biogenesis. The import of cytosolic precursor proteins relies on the Toc- （translocon at the outer chloroplast membrane） and Tic- （translocon at the inner chloroplast membrane） complexes. In Arabidopsis thaliana, precursor recognition is mainly mediated by outer membrane receptors belonging to two gene families： Toc34/33 and Toc159/132/120/90. The role in import and precursor selectivity of these receptors has been intensively studied, but the function of Toc90 still remains unclear. Here, we report the ability of Toc90 to support the import of Toc159 client proteins. We show that the overexpression of Toc90 partially complements the albino knockout of Toc159 and restores photoautotrophic growth. Several lines of evidence including proteome profiling demonstrate the import and accumulation of proteins essential for chloroplast biogenesis and functionality.展开更多
Cytokinins are N^6-substituted adenine derivatives that are involved in the regulation of numerous aspects of plant growth and development. These include the control of cell division, leaf senescence, apical dominance...Cytokinins are N^6-substituted adenine derivatives that are involved in the regulation of numerous aspects of plant growth and development. These include the control of cell division, leaf senescence, apical dominance, sink^source relationship, vascular and embryonic development, and apical meristem activity （Kieber and Schaller, 2014）. In addition, there is increasing evidence for the function of cytokinins in abiotic stress responses （Zwack and Rashotte, 2015）.展开更多
BRI1-ASSOCIATED KINASE 1(BAK1/SERK3)and its closest homolog BAK1-LIKE 1(BKK1/SERK4)are leucine-rich repeat receptor kinases(LRR-RKs)belonging to the SOMATIC EMBRYOGENESIS RECEPTOR KINASE(SERK)family.They act as co-rec...BRI1-ASSOCIATED KINASE 1(BAK1/SERK3)and its closest homolog BAK1-LIKE 1(BKK1/SERK4)are leucine-rich repeat receptor kinases(LRR-RKs)belonging to the SOMATIC EMBRYOGENESIS RECEPTOR KINASE(SERK)family.They act as co-receptors of various other LRR-RKs and participate in multiple signaling events by complexing and transphosphorylating ligand-binding receptors.Initially identified as the brassinosteroid receptor BRASSINOSTEROID INSENSITIVE 1(BRI1)co-receptor,BAK1 also functions in plant immunity by interacting with pattern recognition receptors.Mutations in BAK1 and BKK1 cause severely stunted growth and cell death,characterized as autoimmune cell death.Several factors play a role in this type of cell death,including RKs and components of effector-triggered immunity(ETI)signaling pathways,glycosylation factors,ER quality control components,nuclear trafficking components,ion channels,and Nod-like receptors(NLRs).The Shan lab has recently discovered a novel RK BAK-TO-LIFE 2(BTL2)that interacts with BAK1 and triggers cell death in the absence of BAK1 and BKK1.This RK compensates for the loss of BAK1-mediated pattern-triggered immunity(PTI)by activating phytocytokine-mediated immune and cell death responses.展开更多
Plants rely on a diverse set of small-molecule hormones to regu- late every aspect of their biological processes including develop- ment, growth, and adaptation. Since the discovery of the first plant hormone, auxin, ...Plants rely on a diverse set of small-molecule hormones to regu- late every aspect of their biological processes including develop- ment, growth, and adaptation. Since the discovery of the first plant hormone, auxin, hormones have always been at the frontier of plant biology.展开更多
In order to assess the functional roles of heat stress-induced class B-heat shock factors in Arabidopsis, we investigated T-DNA knockout mutants of AtHsfB1 and AtHsfB2b. Micorarray analysis of double knockout hsfB1/hs...In order to assess the functional roles of heat stress-induced class B-heat shock factors in Arabidopsis, we investigated T-DNA knockout mutants of AtHsfB1 and AtHsfB2b. Micorarray analysis of double knockout hsfB1/hsfB2b plants revealed as strong an up-regulation of the basal mRNA-levels of the defensin genes Pdfl.2a/b in mutant plants. The Pdfexpression was further enhanced by jasmonic acid treatment or infection with the necrotrophic fungus Alternaria brassicicola. The single mutant hsfB2b and the double mutant hsfB 1/B2b were significantly improved in disease resistance after A. brassicicola infection. There was no indication for a direct interaction of Hsf with the promoter of Pdf1.2, which is devoid of perfect HSE consensus Hsf-binding sequences. However, changes in the formation of late HsfA2-dependent HSE binding were detected in hsfB1/B2b plants. This suggests that HsfB1/B2b may interact with class A-Hsf in regulating the shut-off of the heat shock response. The identification of Pdfgenes as targets of Hsf-dependent negative regulation is the first evidence for an interconnection of Hsf in the regulation of biotic and abiotic responses.展开更多
Nitric oxide(NO)is an essential reactive oxygen species and a signal molecule in plants.Although several studies have proposed the occurrence of oxidative NO production,only reductive routes for NO production,such as ...Nitric oxide(NO)is an essential reactive oxygen species and a signal molecule in plants.Although several studies have proposed the occurrence of oxidative NO production,only reductive routes for NO production,such as the nitrate(NO_(3)^(-))-upper-reductase pathway,have been evidenced to date in land plants.However,plants grown axenically with ammonium as the sole source of nitrogen exhibit contents of nitrite and NO3−,evidencing the existence of a metabolic pathway for oxidative production of NO.We hypothesized that oximes,such as indole-3-acetaldoxime(IAOx),a precursor to indole-3-acetic acid,are intermediate oxidation products in NO synthesis.We detected the production of NO from IAOx and other oximes catalyzed by peroxidase(POD)enzyme using both 4-amino-5-methylamino-2′,7′-difluorescein fluorescence and chemiluminescence.Flavins stimulated the reaction,while superoxide dismutase inhibited it.Interestingly,mouse NO synthase can also use IAOx to produce NO at a lower rate than POD.We provided a full mechanism for POD-dependent NO production from IAOx consistent with the experimental data and supported by density functional theory calculations.We showed that the addition of IAOx to extracts from Medicago truncatula increased the in vitro production of NO,while in vivo supplementation of IAOx and other oximes increased the number of lateral roots,as shown for NO donors,and a more than 10-fold increase in IAOx dehydratase expression.Furthermore,we found that in vivo supplementation of IAOx increased NO production in Arabidopsis thaliana wild-type plants,while prx33-34 mutant plants,defective in POD33-34,had reduced production.Our data show that the release of NO by IAOx,as well as its auxinic effect,explain the superroot phenotype.Collectively,our study reveals that plants produce NO utilizing diverse molecules such as oximes,POD,and flavins,which are widely distributed in the plant kingdom,thus introducing a long-awaited oxidative pathway to NO production in plants.This knowledge has essential implications f展开更多
DE-ETIOLATED 1(DET1)and CONSTITUTIVE PHOTOMORPHOGENESIS 1(COP1)are two essential repressors of Arabidopsis photomorphogenesis.These proteins can associate with CULLIN4 to form independent CRL4-based E3 ubiquitin ligas...DE-ETIOLATED 1(DET1)and CONSTITUTIVE PHOTOMORPHOGENESIS 1(COP1)are two essential repressors of Arabidopsis photomorphogenesis.These proteins can associate with CULLIN4 to form independent CRL4-based E3 ubiquitin ligases that mediate the degradation of several photomorphogenic transcription factors,including ELONGATED HYPOCOTYL 5(HY5),thereby controlling multiple gene-regulatory networks.Despite extensive biochemical and genetic analyses of their multi-subunit complexes,the functional links between DET1 and COP1 have long remained elusive.Here,we report that DET1 associates with COP1 in vivo,enhances COP1-HY5 interaction,and promotes COP1 destabilization in a process that dampens HY5 protein abundance.By regulating its accumulation,DET1 avoids HY5 association with hundreds of second-site genomic loci,which are also frequently targeted by the skotomorphogenic transcription factor PHYTOCHROME-INTERACTING FACTOR 3.Accordingly,ectopic HY5 chromatin enrichment favors local gene repression and can trigger fusca-like phenotypes.This study therefore shows that DET1-mediated regulation of COP1 stability tunes down the HY5 cistrome,avoiding hyper-photomorphogenic responses that might compromise plant viability.展开更多
Multilayered defense responses are activated upon pathogen attack.Viruses utilize a number of strategies to maximize the coding capacity of their small genomes and produce viral proteins for infection,including suppre...Multilayered defense responses are activated upon pathogen attack.Viruses utilize a number of strategies to maximize the coding capacity of their small genomes and produce viral proteins for infection,including suppression of host defense.Here,we reveal translation leakage as one of these strategies:two viral effec-tors encoded by tomato golden mosaic virus,chloroplast-localized C4(cC4)and membrane-associated C4(mC4),are translated from two in-frame start codons and function cooperatively to suppress defense.cC4 localizes in chloroplasts,to which it recruits NbPUB4 to induce ubiquitination of the outer membrane;as a result,this organelle is degraded,and chloroplast-mediated defenses are abrogated.However,chloroplast-localized cC4 induces the production of singlet oxygen(^(1)O_(2)),which in turn promotes translo-cation of the ^(1)O_(2) sensor NbMBS1 from the cytosol to the nucleus,where it activates expression of the CERK1 gene.Importantly,an antiviral effect exerted by CERK1 is countered by mC4,localized at the plasma membrane.mC4,like cC4,recruits NbPUB4 and promotes the ubiquitination and subsequent degradation of CERK1,suppressing membrane-based,receptor-like kinase-dependent defenses.Importantly,this translation leakage strategy seems to be conserved in multiple viral species and is related to host range.Thisfinding suggests that stacking of different cellular antiviral responses could be an effective way to abrogate viral infection and engineer sustainable resistance to major crop viral diseases in thefield.展开更多
Protein-protein interactions and the formation of protein complexes play a central role in the regulation of virtually all aspects of life.The identification and characterization of interactions between proteins is th...Protein-protein interactions and the formation of protein complexes play a central role in the regulation of virtually all aspects of life.The identification and characterization of interactions between proteins is the basis of our understanding of protein function and the biological processes they enable.展开更多
Increasing evidence supports the notion that pattern-recognition-receptor-mediated immunity goes beyond plasma membrane-to-nucleus signaling.With the findings of the pathways linking plasma membrane and chloroplasts a...Increasing evidence supports the notion that pattern-recognition-receptor-mediated immunity goes beyond plasma membrane-to-nucleus signaling.With the findings of the pathways linking plasma membrane and chloroplasts and the functions of stromules and perinuclear chloroplast clustering(PCC)in plant defense,chloroplast immunity has emerged as a cornerstone of plant defense and a target of plant pathogens.展开更多
Keeping immune responses in an off state in the absence of a threat is essential to avoid the deleterious consequences of autoimmunity,while their timely activation upon perception of non-self determines the capacity ...Keeping immune responses in an off state in the absence of a threat is essential to avoid the deleterious consequences of autoimmunity,while their timely activation upon perception of non-self determines the capacity of an organism to defend against potential pathogens.In plants,constitutive or untimely activation of immune responses dramatically limits growth and reproduction,highlighting the essential role of negative regulators of immunity for plant fitness.While a number of such regulators have been identified in the past couple of decades,the full complexity of the molecular mechanisms deployed by plants to ensure an appropriately controlled activation of immune responses,as well as their coordination with growth and development,are still largely elusive.展开更多
Positive-sense single-stranded RNA(+ssRNA)viruses,the most abundant viruses of eukaryotes in nature,require the synthesis of negative-sense RNA(-RNA)using their genomic(positive-sense)RNA(+RNA)as a template for replic...Positive-sense single-stranded RNA(+ssRNA)viruses,the most abundant viruses of eukaryotes in nature,require the synthesis of negative-sense RNA(-RNA)using their genomic(positive-sense)RNA(+RNA)as a template for replication.Based on current evidence,viral proteins are translated via viral+RNAs,whereas-RNA is considered to be a viral replication intermediate without coding capacity.Here,we report that plant and animal+ssRNA viruses contain small open reading frames(ORFs)in their-RNA(reverse ORFs[rORFs]).Using turnip mosaic virus(TuMV)as a model for plant+ssRNA viruses,we demonstrate that small proteins encoded by rORFs display specific subcellularlocalizations,and confirm the presence of rORF2 in infected cells through mass spectrometry analysis.The protein encoded by TuMV rORF2 forms punctuate granules that are localized in the perinuclear region and co-localized with viral replication complexes.The rORF2 protein can directly interact with the viral RNA-dependent RNA polymerase,and mutation of rORF2 completely abolishes virus infection,whereas ectopic expression of rORF2 rescues the mutant virus.Furthermore,we show that several rORFs in the-RNA of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)have the ability to suppress type l interferon production and facilitate the infection of ve-sicular stomatitis virus.In addition,we provide evidence that TuMV might utilize internal ribosome entry sites to translate these small rORFs.Taken together,these findings indicate that the-RNA of+ssRNA vi-ruses can also have the coding capacity and that small proteins encoded therein play critical roles in viral infection,revealing a viral proteome larger than previously thought.展开更多
文摘Polyamines （PA）, polyamine oxidases, copper amine oxidases, and nitric oxide （NO） play important roles in physiology and stress responses in plants. NO biosynthesis as a result of catabolism of PA by polyamine oxidases and copper amine oxidases may explain in part PA-mediated responses. Involvement of a copper amine oxidase gene, COPPER AMINE OXIDASEI （CuAO1）, of Arabidopsis was tested for its role in stress responses using the knockouts cuao1-1 and cuaol-2. PA-induced and ABA-induced NO production investigated bY fluorometry and fluorescence microscopy showed that the cuaol-1 and cuaol-2 are impaired in NO production, suggesting a function of CuAO1 in PA and ABA-mediated NO production. Furthermore, we found a PA-dependent increase in protein S-nitrosylation. The addition of PA and ABA also resulted in H2O2 increases, cuao1-1 and cuao1-2 showed less sensitivity to exogenous ABA supplementation during germination, seedling establishment, and root growth inhibition as compared to wild-type. In response to ABA treatment, expression levels of the stress-responsive genes RD29A and ADH1 were significantly lower in the knockouts. These observations characterize cuao1-1 and cuao1-2 as ABA-insensitive mutants. Taken together, our findings extend the ABA signal transduction network to include CuAO1 as one potential contributor to enhanced NO production by ABA.
文摘The polar, sub-cellular localization of PIN auxin efflux carriers determines the direction of intercellular auxin flow, thus defining the spatial aspect of auxin signalling. Dynamic, transcytosis-like relocalizations of PIN proteins occur in response to external and internal signals, integrating these signals into changes in auxin distribution. Here, we examine the cellular and molecular mechanisms of polar PIN delivery and transcytosis. The mechanisms of the ARF-GEF-dependent polar targeting and transcytosis are well conserved and show little variations among diverse Arabidopsis ecotypes consistent with their fundamental importance in regulating plant development. At the cellular level, we refine previous findings on the role of the actin cytoskeleton in apical and basal PIN targeting, and identify a previously unknown role for microtubules, specifically in basal targeting. PIN protein delivery to different sides of the cell is mediated by ARFdependent trafficking with a previously unknown complex level of distinct ARF-GEF vesicle trafficking regulators. Our data suggest that alternative recruitment of PIN proteins by these distinct pathways can account for cell type- and cargo-specific aspects of polar targeting, as well as for polarity changes in response to different signals. The resulting dynamic PIN positioning to different sides of cells defines a three-dimensional pattern of auxin fluxes within plant tissues.
文摘As the first and rate-limiting enzyme of proline degradation, PROLINE DEHYDROGENASE1 （PDH1） is tightly regulated during plant stress responses, including induction under hypoosmolarity and repression under water deficit. The plant receptor histidine kinases AHKs, elements of the two-component system （TCS） in Arabidopsis thaliana, are pro- posed to function in water stress responses by regulating different stress-responsive genes. However, little information is available concerning AHK phosphorelay-mediated downstream signaling. Here we show that the Arabidopsis type-B response regulator 18 （ARR18） functions as a positive osmotic stress response regulator in Arabidopsis seeds and affects the activity of the PDH1 promoter, known to be controlled by C-group bZlP transcription factors. Moreover, direct physical interaction of ARR18 with bZIP63 was identified and shown to be dependent on phosphorylation of the conserved aspar- tate residue in the ARR18 receiver domain. We further show that bZlP63 itself functions as a negative regulator of seed germination upon osmotic stress. Using reporter gene assays in protoplasts, we demonstrated that ARR18 interaction negatively interferes with the transcriptional activity of bZIP63 on the PDH1 promoter. Our findings provide new insight into the function of ARR18 and bZIP63 as antagonistic regulators of gene expression in Arabidopsis.
文摘High-temperature stress, like any abiotic stress, impairs the physiology and development of plants, including the stages of seed setting and ripening. We used the Affymetrix 22K Barley1 GeneChip microarray to investigate the response of developing barley （Hordeum vulgare） seeds, termed caryopses, after 0.5, 3, and 6 h of heat stress exposure; 958 induced and 1122 repressed genes exhibited spatial and temporal expression patterns that provide a detailed insight into the caryopses＇ early heat stress responses. Down-regulation of genes related to storage compound biosynthesis and cell growth provides evidence for a rapid impairment of the caryopsis＇ development. Increased levels of sugars and amino acids were indicative for both production of compatible solutes and feedback-induced accumulation of substrates for storage compound biosynthesis. Metadata analysis identified embryo and endosperm as primary locations of heat stress responses, indicating a strong impact of short-term heat stress on central developmental functions of the caryopsis. A comparison with heat stress responses in Arabidopsis shoots and drought stress responses in barley caryopses identified both conserved and presumably heat- and caryopsis-specific stress-responsive genes. Summarized, our data provide an important basis for further investigation of gene functions in order to aid an improved heat tolerance and reduced losses of yield in barley as a model for cereal crops.
文摘Besides providing protection against mechanical damage to the root tip, the root cap is involved in the perception and processing of diverse external and internal stimuli resulting in altered growth and development, The transduction of these stimuli includes hormonal signaling pathways such as those of auxin, ethylene and cytokinin. Here, we show that the root cap is essential for the ethylene-induced regulation of elongation growth and root hair formation in maize. Exogenously applied ethylene is no longer able to inhibit elongation growth when the root cap has been surgically removed prior to hormone treatment. Reconstitution of the cap positively correlates with the developing capacity of the roots to respond to ethylene again. In contrast, the removal of the root cap does not per se affect growth inhibition controlled by auxin and cytokinin. Furthermore, our semi-quantitative RT-PCR results support earlier findings that the maize root cap is a site of high gene expression activity with respect to sensing and responding to hormones such as ethylene, From these data, we propose a novel function of the root cap which is the establishment of competence to respond to ethylene in the distal zones of the root.
文摘In order to analyze the signaling function of hydrogen peroxide （H202） production in senescence in more detail, we manipulated intracellular H202 levels in Arabidopsis thaliala （L.） Heynh by using the hydrogen- peroxide-sensitive part of the Escherichia coil transcription regulator OxyR, which was directed to the cytoplasm as well as into the peroxisomes. H202 levels were lowered and senescence was delayed in both transgenic lines, but OxyR was found to be more effective in the cytoplasm. To transfer this knowledge to crop plants, we analyzed oilseed rape plants Brassica napus L. cv. Mozart for H20~ and its scavenging enzymes catalase （CAT） and ascorbate peroxidase （APX） during leaf and plant development. H202 levels were found to increase during bolting and flowering time, but no increase could be observed in the very late stages of senescence. With increasing H202 levels, CAT and APX activities declined, so it is likely that similar mechanisms are used in oilseed rape and Arabidopsis to control H202 levels. Under elevated CO2 conditions, oilseed rape senescence was accelerated and coincided with an earlier increase in H202 levels, indicating that H202 may be one of the signals to inducing senescence in a broader range of Brassicaceae.
文摘Spectro-microscopy, a combination of fluorescence microscopy with spatially resolved spectroscopic techni- ques, provides new and exciting tools for functional cell biology in living organisms. This review focuses on recent devel- opments in spectro-microscopic applications for the investigation of living plant cells in their native tissue context. The application of spectro-microscopic methods led to the recent discovery of a fast signal response pathway for the brassi- nosteroide receptor BRI1 in the plasma membrane of living plant cells. Moreover, the competence of different plant cell types to respond to environmental or endogenous stimuli was determined in vivo by correlation analysis of different optical and spectroscopic readouts such as fluorescence lifetime （FLT）. Furthermore, a new spectro-microscopic technique, fluorescence intensity decay shape analysis microscopy （FIDSAM）, has been developed. FIDSAM is capable of imaging low- expressed fluorophore-tagged proteins at high spatial resolution and precludes the misinterpretation of autofluorescence artifacts. In addition, FIDSAM provides a very effective and sensitive tool on the basis of F6rster resonance energy transfer （FRET） for the qualitative and quantitative determination of protein-protein interaction. Finally, we report on the quan- titative analysis of the photosystem I and II （PSI/PSII） ratio in the chloroplasts of living Arabidopsis plants at room tem- perature, using high-resolution, spatially resolved fluorescence spectroscopy. With this technique, it was not only possible to measure PSI/PSII ratios, but also to demonstrate the differential competence of wild-type and carbohydrate-deficient plants to adapt the PSI/PSII ratio to different light conditions. In summary, the information content of standard microscopic images is extended by several dimensions by the use of spectro-microscopic approaches. Therefore, novel cell physiolog- ical and molecular topics can be addressed and valuable insights into molecular and subcellula
基金support from tho Europoan Rosearch Council to R.G.(ERC Consolidator Grant"bi-BL OCK"ID 646644,ERC Proof of Concept Grant"TnVolve"ID 957547).
文摘Seed size critically affects grain yield of crops and hence represents a key breeding target.The develop-ment of embryo-nourishing endosperm is a key driver of seed expansion.We here report unexpected dual roles of the transcription factor EIN3 in regulating seed size.These EIN3 functions have remained largely undiscovered because they oppose each other.Capitalizing on the analysis of multiple ethylene biosynthesis mutants,we demonstrate that EIN3 represses endosperm and seed development in a pathway regulated by ethylene.We,in addition,provide evidence that EIN3-mediated synergid nucleus disintegration promotes endosperm expansion.Interestingly,synergid nucleus disintegration is not affected in various ethylene biosynthesis mutants,suggesting that this promoting function of EIN3 is inde-pendent of ethylene.Whereas the growth-inhibitory ethylene-dependent EIN3 action appears to be encoded by sporophytic tissue,the growth-promoting role of EIN3 is induced by fertilization,revealing a generation conflict that converges toward the key signaling component EIN3.
文摘Successful import of hundreds of nucleus-encoded proteins is essential for chloroplast biogenesis. The import of cytosolic precursor proteins relies on the Toc- （translocon at the outer chloroplast membrane） and Tic- （translocon at the inner chloroplast membrane） complexes. In Arabidopsis thaliana, precursor recognition is mainly mediated by outer membrane receptors belonging to two gene families： Toc34/33 and Toc159/132/120/90. The role in import and precursor selectivity of these receptors has been intensively studied, but the function of Toc90 still remains unclear. Here, we report the ability of Toc90 to support the import of Toc159 client proteins. We show that the overexpression of Toc90 partially complements the albino knockout of Toc159 and restores photoautotrophic growth. Several lines of evidence including proteome profiling demonstrate the import and accumulation of proteins essential for chloroplast biogenesis and functionality.
文摘Cytokinins are N^6-substituted adenine derivatives that are involved in the regulation of numerous aspects of plant growth and development. These include the control of cell division, leaf senescence, apical dominance, sink^source relationship, vascular and embryonic development, and apical meristem activity （Kieber and Schaller, 2014）. In addition, there is increasing evidence for the function of cytokinins in abiotic stress responses （Zwack and Rashotte, 2015）.
基金supported by the SFB1101 D03 and the TRR356 B02,funded by the German science foundation DFG.
文摘BRI1-ASSOCIATED KINASE 1(BAK1/SERK3)and its closest homolog BAK1-LIKE 1(BKK1/SERK4)are leucine-rich repeat receptor kinases(LRR-RKs)belonging to the SOMATIC EMBRYOGENESIS RECEPTOR KINASE(SERK)family.They act as co-receptors of various other LRR-RKs and participate in multiple signaling events by complexing and transphosphorylating ligand-binding receptors.Initially identified as the brassinosteroid receptor BRASSINOSTEROID INSENSITIVE 1(BRI1)co-receptor,BAK1 also functions in plant immunity by interacting with pattern recognition receptors.Mutations in BAK1 and BKK1 cause severely stunted growth and cell death,characterized as autoimmune cell death.Several factors play a role in this type of cell death,including RKs and components of effector-triggered immunity(ETI)signaling pathways,glycosylation factors,ER quality control components,nuclear trafficking components,ion channels,and Nod-like receptors(NLRs).The Shan lab has recently discovered a novel RK BAK-TO-LIFE 2(BTL2)that interacts with BAK1 and triggers cell death in the absence of BAK1 and BKK1.This RK compensates for the loss of BAK1-mediated pattern-triggered immunity(PTI)by activating phytocytokine-mediated immune and cell death responses.
文摘Plants rely on a diverse set of small-molecule hormones to regu- late every aspect of their biological processes including develop- ment, growth, and adaptation. Since the discovery of the first plant hormone, auxin, hormones have always been at the frontier of plant biology.
文摘In order to assess the functional roles of heat stress-induced class B-heat shock factors in Arabidopsis, we investigated T-DNA knockout mutants of AtHsfB1 and AtHsfB2b. Micorarray analysis of double knockout hsfB1/hsfB2b plants revealed as strong an up-regulation of the basal mRNA-levels of the defensin genes Pdfl.2a/b in mutant plants. The Pdfexpression was further enhanced by jasmonic acid treatment or infection with the necrotrophic fungus Alternaria brassicicola. The single mutant hsfB2b and the double mutant hsfB 1/B2b were significantly improved in disease resistance after A. brassicicola infection. There was no indication for a direct interaction of Hsf with the promoter of Pdf1.2, which is devoid of perfect HSE consensus Hsf-binding sequences. However, changes in the formation of late HsfA2-dependent HSE binding were detected in hsfB1/B2b plants. This suggests that HsfB1/B2b may interact with class A-Hsf in regulating the shut-off of the heat shock response. The identification of Pdfgenes as targets of Hsf-dependent negative regulation is the first evidence for an interconnection of Hsf in the regulation of biotic and abiotic responses.
基金supported by grants AGL2014-52396,AGL2017-86293-P,and PID2022-142968NB-I00 from MCIN/AEI/10.13039/501100011033/FEDER,UE,and a grant from the Public University of Navarre(PID-2020-117703GB-I00)(to J.F.M.)and the UPV/EHU-GV IT-1018-16 program(Basque Government)(to R.E.).M.U.is a recipient of a predoctoral fellowship from the Government of Navarre,Spain.J.B.and P.L.-G.have received pre-doctoral fellowships from the Public University of Navarre,Spain.P.L.-G is currently financed by a postdoctoral contract funded by the Spanish National Research Council(20224AT017).J.B.is also a recipient of the"Requalification of the Spanish University System for 2021-2023,Public University of Navarra"fellowship,funded by the European Union-Next Generation(EU).
文摘Nitric oxide(NO)is an essential reactive oxygen species and a signal molecule in plants.Although several studies have proposed the occurrence of oxidative NO production,only reductive routes for NO production,such as the nitrate(NO_(3)^(-))-upper-reductase pathway,have been evidenced to date in land plants.However,plants grown axenically with ammonium as the sole source of nitrogen exhibit contents of nitrite and NO3−,evidencing the existence of a metabolic pathway for oxidative production of NO.We hypothesized that oximes,such as indole-3-acetaldoxime(IAOx),a precursor to indole-3-acetic acid,are intermediate oxidation products in NO synthesis.We detected the production of NO from IAOx and other oximes catalyzed by peroxidase(POD)enzyme using both 4-amino-5-methylamino-2′,7′-difluorescein fluorescence and chemiluminescence.Flavins stimulated the reaction,while superoxide dismutase inhibited it.Interestingly,mouse NO synthase can also use IAOx to produce NO at a lower rate than POD.We provided a full mechanism for POD-dependent NO production from IAOx consistent with the experimental data and supported by density functional theory calculations.We showed that the addition of IAOx to extracts from Medicago truncatula increased the in vitro production of NO,while in vivo supplementation of IAOx and other oximes increased the number of lateral roots,as shown for NO donors,and a more than 10-fold increase in IAOx dehydratase expression.Furthermore,we found that in vivo supplementation of IAOx increased NO production in Arabidopsis thaliana wild-type plants,while prx33-34 mutant plants,defective in POD33-34,had reduced production.Our data show that the release of NO by IAOx,as well as its auxinic effect,explain the superroot phenotype.Collectively,our study reveals that plants produce NO utilizing diverse molecules such as oximes,POD,and flavins,which are widely distributed in the plant kingdom,thus introducing a long-awaited oxidative pathway to NO production in plants.This knowledge has essential implications f
基金supported by a Ramon y Cajal(RYC-2014-16308)grant funded by the Ministerio de Economfa y Competitividad to S.F.Work by S.F.in F.B.’s lab was supported by the COST Action CA16212 INDEPTH(European Union)funded by the Agencia Estatal de Investigacion/Fondo Europeo de Desarollo Regional/European Union(BIO2016-80551-R and PID2019-105495GB-I00).+2 种基金supported by CNRS EPIPLANT Action(France)and funded by Agence Nationale de la Recherche grants ANR-10-LABX-54,ANR-18-CE13-0004-01,ANR-17-CE12-0026-02(France)by Velux Stiftung(Switzerland).B.G.G.is funded by President's International Fellowship Initiative postdoctoral fellowship(no.2020PB0082)the Chinese Academy of Sciences,and is the recipient of a Talent-Introduction grant(Chinese Postdoctoral International Exchange Program).
文摘DE-ETIOLATED 1(DET1)and CONSTITUTIVE PHOTOMORPHOGENESIS 1(COP1)are two essential repressors of Arabidopsis photomorphogenesis.These proteins can associate with CULLIN4 to form independent CRL4-based E3 ubiquitin ligases that mediate the degradation of several photomorphogenic transcription factors,including ELONGATED HYPOCOTYL 5(HY5),thereby controlling multiple gene-regulatory networks.Despite extensive biochemical and genetic analyses of their multi-subunit complexes,the functional links between DET1 and COP1 have long remained elusive.Here,we report that DET1 associates with COP1 in vivo,enhances COP1-HY5 interaction,and promotes COP1 destabilization in a process that dampens HY5 protein abundance.By regulating its accumulation,DET1 avoids HY5 association with hundreds of second-site genomic loci,which are also frequently targeted by the skotomorphogenic transcription factor PHYTOCHROME-INTERACTING FACTOR 3.Accordingly,ectopic HY5 chromatin enrichment favors local gene repression and can trigger fusca-like phenotypes.This study therefore shows that DET1-mediated regulation of COP1 stability tunes down the HY5 cistrome,avoiding hyper-photomorphogenic responses that might compromise plant viability.
基金funded by grants from the National Natural Science Foundation of China (31930089)the National Key Research and Development Program of China (2021YFD1400400)the Yunnan Zhouxueping Expert Workstation (No.202205AF150047).
文摘Multilayered defense responses are activated upon pathogen attack.Viruses utilize a number of strategies to maximize the coding capacity of their small genomes and produce viral proteins for infection,including suppression of host defense.Here,we reveal translation leakage as one of these strategies:two viral effec-tors encoded by tomato golden mosaic virus,chloroplast-localized C4(cC4)and membrane-associated C4(mC4),are translated from two in-frame start codons and function cooperatively to suppress defense.cC4 localizes in chloroplasts,to which it recruits NbPUB4 to induce ubiquitination of the outer membrane;as a result,this organelle is degraded,and chloroplast-mediated defenses are abrogated.However,chloroplast-localized cC4 induces the production of singlet oxygen(^(1)O_(2)),which in turn promotes translo-cation of the ^(1)O_(2) sensor NbMBS1 from the cytosol to the nucleus,where it activates expression of the CERK1 gene.Importantly,an antiviral effect exerted by CERK1 is countered by mC4,localized at the plasma membrane.mC4,like cC4,recruits NbPUB4 and promotes the ubiquitination and subsequent degradation of CERK1,suppressing membrane-based,receptor-like kinase-dependent defenses.Importantly,this translation leakage strategy seems to be conserved in multiple viral species and is related to host range.Thisfinding suggests that stacking of different cellular antiviral responses could be an effective way to abrogate viral infection and engineer sustainable resistance to major crop viral diseases in thefield.
基金partially funded by the Excellence Strategy of the German Federal and State Governments,the ERC-COG Gem Omics(101044142)the Deutsche Forschungsgemeinschaft(DFG,German Research Foundation)(Project Numbers LO 2314/1-1 and SBF 1101/3,C08)。
文摘Protein-protein interactions and the formation of protein complexes play a central role in the regulation of virtually all aspects of life.The identification and characterization of interactions between proteins is the basis of our understanding of protein function and the biological processes they enable.
基金supported by grants from the National Key Research and Development Program of China(2021YFD1400400)the Natural Science Foundation of China(32320103010,32172385,and 31930089).
文摘Increasing evidence supports the notion that pattern-recognition-receptor-mediated immunity goes beyond plasma membrane-to-nucleus signaling.With the findings of the pathways linking plasma membrane and chloroplasts and the functions of stromules and perinuclear chloroplast clustering(PCC)in plant defense,chloroplast immunity has emerged as a cornerstone of plant defense and a target of plant pathogens.
文摘Keeping immune responses in an off state in the absence of a threat is essential to avoid the deleterious consequences of autoimmunity,while their timely activation upon perception of non-self determines the capacity of an organism to defend against potential pathogens.In plants,constitutive or untimely activation of immune responses dramatically limits growth and reproduction,highlighting the essential role of negative regulators of immunity for plant fitness.While a number of such regulators have been identified in the past couple of decades,the full complexity of the molecular mechanisms deployed by plants to ensure an appropriately controlled activation of immune responses,as well as their coordination with growth and development,are still largely elusive.
基金funded by the National Key Research and Development Program of China(2021YFD1400400)to F.L.the National Natural Science Foundation of China(31930089 and 31972244)to X.Z.and F.L.+2 种基金a startup grant for High-level Talents of Fujian Medical University(XRCZX2019019)the Natural Science Foundation of Fujan Province,China(2020J01604)to Q.S.Work in the R.L.-D.lab is partially funded by the ERC-COG grant GemOmics(101044142)to R.L.-D.
文摘Positive-sense single-stranded RNA(+ssRNA)viruses,the most abundant viruses of eukaryotes in nature,require the synthesis of negative-sense RNA(-RNA)using their genomic(positive-sense)RNA(+RNA)as a template for replication.Based on current evidence,viral proteins are translated via viral+RNAs,whereas-RNA is considered to be a viral replication intermediate without coding capacity.Here,we report that plant and animal+ssRNA viruses contain small open reading frames(ORFs)in their-RNA(reverse ORFs[rORFs]).Using turnip mosaic virus(TuMV)as a model for plant+ssRNA viruses,we demonstrate that small proteins encoded by rORFs display specific subcellularlocalizations,and confirm the presence of rORF2 in infected cells through mass spectrometry analysis.The protein encoded by TuMV rORF2 forms punctuate granules that are localized in the perinuclear region and co-localized with viral replication complexes.The rORF2 protein can directly interact with the viral RNA-dependent RNA polymerase,and mutation of rORF2 completely abolishes virus infection,whereas ectopic expression of rORF2 rescues the mutant virus.Furthermore,we show that several rORFs in the-RNA of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)have the ability to suppress type l interferon production and facilitate the infection of ve-sicular stomatitis virus.In addition,we provide evidence that TuMV might utilize internal ribosome entry sites to translate these small rORFs.Taken together,these findings indicate that the-RNA of+ssRNA vi-ruses can also have the coding capacity and that small proteins encoded therein play critical roles in viral infection,revealing a viral proteome larger than previously thought.
