气候变化对海藻龙须菜生长与光合作用耐热特性的影响

为探讨大气CO2升高和温室效应对龙须菜生长及生理生化特性的影响,在4种条件下培养龙须菜:1)对照组(390μL/L CO2+20℃),2)CO2升高组(700μL/L CO2+20℃),3)温度升高组(390μL/L CO2+24℃),4)温室效应组(700μL/L CO2+24℃),测定藻体生长和生化组分以及高温胁迫下的最大光化学量子产量(Fv/Fm)和光能利用效率(α)、光合速率(Pn)和呼吸速率(Rd)。结果表明,CO2升高、温度升高以及温室效应均促进龙须菜的生长,温室效应下的促进作用更明显。温室效应使龙须菜具较高的Pn和Rd以及较低的可溶性蛋白(SP)和可溶性碳水化合物(SC)含量。高浓度CO2对叶绿素(Chl a)和类胡萝卜素(Car)含量没有显著影响,而高温使其上升;藻红蛋白(PE)和藻蓝蛋白(PC)含量不受CO2浓度和温度的影响。龙须菜Fv/Fm、α、Pn和Rd值,在32℃处理3 h后略有上升,在36℃处理3 h后下降,而在40℃处理20 min后降到极低水平。正常温度(20℃)生长的龙须菜最高耐受温度在32—36℃之间,而较高温(24℃)生长的龙须菜在36—40℃之间;生长温度对光合作用和呼吸作用耐热性能的影响比CO2浓度的影响更大;而温室效应生长条件下的龙须菜光合作用表现出更突出的耐热性能。     

大气CO2升高和紫外辐射相互作用对羊栖菜生理特性的影响

为了研究经济海藻羊栖菜对大气CO2浓度增加与紫外辐射(UVR)相互作用的响应,设置两个CO2浓度(380μL/L和800μL/L)以及两种辐射处理,即PAR处理(滤除UV-A、UV-B,藻体仅接受可见光,400—700nm)和PAB处理(全波长辐射280—700nm)培养海藻,探讨了羊栖菜生长、光合作用、呼吸作用、光合色素含量、可溶性糖和蛋白以及硝酸还原酶活性的变化情况。结果表明高浓度CO2显著提高羊栖菜藻体的相对生长速率,并且紫外辐射的负面效应在高CO2处理下表现不显著。高CO2降低了藻体的光合作用速率,而UVR的负面效应和生长体现为一致性,但是羊栖菜的呼吸作用没有受到环境变化的明显影响。羊栖菜的光合色素叶绿素a和类胡萝卜素在高浓度CO2处理下明显降低,而UVR没有明显影响。环境因子对羊栖菜的可溶性糖没有影响,但是在高CO2和全波长辐射处理下,藻体可溶性蛋白的含量显著增加。同时高CO2明显提高了硝酸还原酶的活性,并且仅在高浓度CO2处理下藻体中UVR对其活性有抑制作用。CO2和UVR对羊栖菜的大多数生理特性存在明显的交互作用,在未来CO2浓度进一步增加的情况下,UVR的负面效应将会得到一定程度的缓解,这样有利于羊栖菜在养殖海区获得更高的产量。     

大气CO_2浓度升高对不同氮生长条件下的两种大型海藻光合作用的影响

以龙须菜和蛎菜两种大型海藻为试验材料,在室外自然条件下培养,设置常规CO2浓度(390μL·L-1)和高CO2浓度(800μL·L-1)空气两种通气条件、以及10和150μmol·L-1两种氮营养盐供应水平,以探讨大气CO2浓度升高对不同氮营养盐生长条件下海藻生长与光合特性的影响。结果表明:不管是高氮还是低氮的培养条件,大气CO2浓度升高都能促进这两种海藻的生长;在龙须菜中,这种生长促进作用在低氮培养条件下更高,而在蛎菜中这种生长促进作用在高氮培养条件下较高;短期海水中高CO2浓度能促进蛎菜和龙须菜的光合作用;长期高CO2浓度生长环境依然能维持龙须菜高的光合作用,但抑制蛎菜的光合作用。另外,不管培养液中CO2供应条件如何,氮加富均促进了龙须菜和蛎菜这两种海藻的光全和呼吸作用。     

发菜(Nostoc flagelliforme)培养条件的研究

研究了光照强度、日供水次数、CO2浓度和培养基成分对发菜生长的影响。结果显示,中度光强(114μmol.m-2.s-1)下发菜生长最快;发菜的生长基本同供水次数成正相关系;CO2浓度的升高并没有显著促进发菜生长,低光条件下(57μmol.m-2.s-1),高浓度的CO2(2800μL/L)抑制了发菜的生长;用BG11培养的发菜生物量的增长显著高于用BG110培养的;BG11培养基中K+和CO32-的缺失并没有显著影响发菜的生长。     

在海洋酸化背景下温度变化对大型绿藻缘管浒苔光合生理特性的影响

为了探讨在未来海洋酸化背景下温度变化对大型海藻的影响,选取了绿潮藻缘管浒苔(Ulva linza)作为实验材料。实验设置2个CO2浓度,分别为正常空气水平(LC:390μL·L-1)和高CO2水平(HC:1000μL·L-1),在20℃下从缘管浒苔的幼苗开始培养,等成体后转入不同的温度梯度(5、10、15、20、25和30℃)下测定藻体的光合作用。结果表明:海洋酸化显著提高了缘管浒苔的相对生长速率,虽然藻体光合作用速率没有明显变化,但藻体的呼吸作用和硝酸还原酶能力显著增加;对于光合色素来说,高CO2显著降低了叶绿素a和叶绿素b的含量;温度变化在不同CO2处理下表现为不同的效应;长期生长在20℃的缘管浒苔,当环境温度降低时,其最大光合作用速率Pmax、呼吸作用Rd、光饱和点Ik和光补偿点Ic都随之降低,而光合效率α则是先升高再降低,其中高CO2提高了藻体的α、Rd和Ic;而当温度增加时,缘管浒苔的Pmax和Ik都随之升高,α、Rd和Ic都有下降的趋势,高温下高CO2的α和Rd较高,光补偿点较低。相较于正常环境来说,海洋酸化下的缘管浒苔对温度的骤变具有更好的适应能力。     

液体悬浮培养条件下发菜细胞的光合速率与呼吸速率

用液相氧电极测定离体悬浮生长发菜细胞的光合速率和呼吸速率的结果表明,发菜细胞的光补偿点为15￣16μmol·m-2·s-1,光饱和点为90μmol·m-2·s-1,光抑制点为190μmol·m-2·s-1。25℃下发菜细胞光合速率最高,呼吸速率则在10￣50℃范围内随温度升高而增强。发菜细胞光合作用的最适pH值为7.0￣7.5,呼吸作用最适pH值为9.0。BG110无氮培养基中添加30mmol·L-1NaNO3,发菜细胞的光合速率增加约20%。培养基中Na2HPO4浓度为1.75mmol·L-1时,细胞光合速率和呼吸速率最大,随后保持稳定。钾盐浓度变化对发菜细胞光合速率和呼吸速率的影响不显著。     

从光合作用特性看铜绿微囊藻(Microcystis aeruginosa)的竞争优势

通过测定净光合放氧速率,研究了温度、光照和pH对铜绿微囊藻(M icrocystis aeruginosa)和玫瑰拟衣藻(Chlorom onas rosae)光合作用的影响。两种藻的光合放氧速率都随着温度的升高而加快,在10~35℃范围内,铜绿微囊藻净光合放氧速率随温度升高而直线上升,其最适温度高于35℃,而当温度高于30℃后玫瑰拟衣藻的净光合放氧速率迅速下降;两种微藻的光合放氧速率-光强变化曲线有所不同,铜绿微囊藻光饱和点在500μmol.m-2.s-1附近,光强达到900μmol.m-2.s-1时仍无光抑制现象发生,玫瑰拟衣藻光饱和点在630μmol.m-2.s-1附近,当光强进一步升高,光合放氧速率开始下降;铜绿微囊藻最适pH值是10.0,在pH值6.5~11.5范围内,光合放氧都很活跃,变化幅度不大,玫瑰拟衣藻最适pH值7.0,偏酸或偏碱光合放氧都迅速地下降,pH高于10.0出现了负值。比较两种藻的光合作用特性,铜绿微囊藻光合作用具有3个特点:(1)适应温度范围宽,对高温具有良好的适应性,并且光合作用随温度的升高显著提高;(2)光饱和点低,光合作用活性高,能在弱光环境中高效地进行光合作用,并且抗强光伤害;(3)对pH变化具有超强的适应能力,在中性和碱性环境中,都能进行活跃的光合作用。铜绿微囊藻在光能利用、温度和pH适应性方面的特点,可以使其快速生长繁殖,积累大量的生物量,在与其它藻类的竞争中占据显著的优势。     

光强和硝态氮水平对铜藻(Sargassum horneri)生长和光合生理特性的影响

以金潮种铜藻(Sargassum horneri)为对象,探讨了不同硝态氮水平下光照强度对其生长和光合生理特性的影响,以期为金潮暴发机制的研究提供一定的参考。实验设置了高(100μmol·L-1)、低(自然海水,50μmol·L-1) 2个硝态氮浓度和高(150μmol photons·m-2·s-1)、中(60μmol photons·m-2·s-1)、低(10μmol photons·m-2·s-1)三个光照水平,检测了藻体在不同硝态氮浓度和光照条件下适应培养7 d后生长、色素含量、可溶性糖含量、可溶性蛋白含量、氮吸收速率及最大光合放氧速率(Pmax)等生理指标的变化。结果表明:氮浓度相同时,铜藻的生长速率、可溶性糖含量、可溶性蛋白含量、氮吸收速率、Pmax均随光照强度的增加而增加,而色素含量随着光照强度的增加而降低;低氮条件下,除类胡萝卜素外,以上指标在高光处理下均显著高于低光处理(P0.05);高氮条件下,除可溶性糖外,以上指标在高光条件下也均显著高于低光处理(P0.05);同一光照强度下,色素及可溶性蛋白含量、氮吸收速率及Pmax随氮浓度的增加而增加;可溶性糖含量随着硝态氮浓度的增加而减少;且在3个光照条件下,高氮处理下藻体的Pmax均显著高于低氮处理。总之,不同光照和氮浓度处理7 d后,高光、高氮适应后的藻体表现出较高的生长和光合作用潜能。铜藻断裂后漂浮于海面极易获得较高光强条件,而富营养海水中的高氮条件会刺激铜藻的光合潜能而使其加速生长,进而加剧铜藻金潮的暴发。     

米瓦罐的光合特性研究

采用美国LI-COR生产的LI-6400便携式光合系统研究了米瓦罐的光合特性。结果表明,米瓦罐光 合速率的日变化呈单峰曲线,上午10:00时光合速率达到最大值;在大气CO2浓度下,米瓦罐的光饱和点为 1 800μmol·m-2·s-1,光补偿点为30μmol·m-2·s-1;在光饱和点的光强下,米瓦罐的CO2饱和点为1 200 μmolCO2·mol-1,CO2补偿点为40μmolCO2·mol-1。     

不同培养条件对枝鞘藻生长、虾青素和油脂积累的影响

以丝状绿藻枝鞘藻(Oedocladium sp.)为实验材料,研究在100、300μmol·m-2·s-1和双侧300μmol·m-2·s-13种光强以及1、3、9、18 mmol/L 4种初始氮浓度下,两步法培养(第12 d时实验组分别更换为无氮培养基及加盐培养基)对枝鞘藻生长、油脂和虾青素积累的影响。结果显示:枝鞘藻最大生物量在双侧300μmol·m-2·s-1光强,18 mmol/L初始氮浓度更换为无氮培养基的条件下达到,为9.61 g/L;最高虾青素含量和最高油脂含量在双侧300μmol·m-2·s-1光强,3 mmol/L初始氮浓度更换为加盐培养基条件下达到,分别达到干重的1.62%和51.19%。研究结果表明高光条件有利于枝鞘藻的生长,双侧高光条件下低氮浓度更换为加盐培养基最有利于枝鞘藻虾青素和油脂的积累。     

Thermodynamic and kinetic characterisation of individual haems in multicentre cytochromes c3

The characterisation of individual centres in multihaem proteins is difficult due to the similarities in the redox and spectroscopic properties of the centres. NMR has been used successfully to distinguish redox centres and allow the determination of the microscopic thermodynamic parameters in several multihaem cytochromes c₃ isolated from different sulphate-reducing bacteria. In this article we show that it is also possible to discriminate the kinetic properties of individual centres in multihaem proteins, if the complete microscopic thermodynamic characterisation is available and the system displays fast intramolecular equilibration in the time scale of the kinetic experiment. The deconvolution of the kinetic traces using a model of thermodynamic control provides a reference rate constant for each haem that does not depend on driving force and can be related to structural factors. The thermodynamic characterisation of three tetrahaem cytochromes and their kinetics of reduction by sodium dithionite are reported in this paper. Thermodynamic and kinetic data were fitted simultaneously to a model to obtain microscopic reduction potentials, haem-haem and haem-proton interacting potentials, and reference rate constants for the haems. The kinetic information obtained for these cytochromes and recently published data for other multihaem cytochromes is discussed with respect to the structural factors that determine the reference rates. The accessibility for the reducing agent seems to play an important role in controlling the kinetic rates, although is clearly not the only factor.     

Electron transfer kinetics between soluble modules of Paracoccus denitrificans cytochrome c1 and its physiological redox partners

The transient electron transfer (ET) interactions between cytochrome c₁ of the bc₁-complex from Paracoccus denitrificans and its physiological redox partners cytochrome c₅₅₂ and cytochrome c₅₅₀ have been characterized functionally by stopped-flow spectroscopy. Two different soluble fragments of cytochrome c₁ were generated and used together with a soluble cytochrome c₅₅₂ module as a model system for interprotein ET reactions. Both c₁ fragments lack the membrane anchor; the c₁ core fragment (c_1CF) consists of only the hydrophilic heme-carrying domain, whereas the c₁ acidic fragment (c_1AF) additionally contains the acidic domain unique to P. denitrificans. In order to determine the ionic strength dependencies of the ET rate constants, an optimized stopped-flow protocol was developed to overcome problems of spectral overlap, heme autoxidation and the prevalent non-pseudo first order conditions. Cytochrome c₁ reveals fast bimolecular rate constants (10⁷ to 10⁸ M^− 1 s^− 1) for the ET reaction with its physiological substrates c₅₅₂ and c₅₅₀, thus approaching the limit of a diffusion-controlled process, with 2 to 3 effective charges of opposite sign contributing to these interactions. No direct involvement of the N-terminal acidic c₁-domain in electrostatically attracting its substrates could be detected. However, a slight preference for cytochrome c₅₅₀ over c₅₅₂ reacting with cyochrome c₁ was found and attributed to the different functions of both cytochromes in the respiratory chain of P. denitrificans.     

Arrest of oogenesis in the bug Rhodnius prolixus challenged with the fungus Aspergillus niger is mediated by immune response-derived PGE2

In this work we characterized the immune response of the insect Rhodnius prolixus to a direct injection into the hemocoel of the non-entomopathogenic fungus Aspergillus niger, and evaluated its consequences on host oogenesis. These animals were able to respond by mounting effective cellular and humoral responses to this fungus; these responses were shown, however, to have reproductive fitness costs, as the number of eggs laid per female was significantly reduced. The disturbance of egg formation during infectious process correlated with an elevation in the titer of hemolymph prostaglandin E₂ 48 h post-challenge. Administration of Zymosan A as an immunogenic non-infectious challenge produced similar effects on phenoloxidase and prophenoloxidase activities, oocyte development and prostaglandin E₂ titer, precluding the hypothesis of an effect mediated by fungal metabolites in animals challenged with fungus. Ovaries at 48 h post-challenge showed absence of vitellogenic ovarian follicles, and the in vivo administration of prostaglandin E₂ or its receptor agonist misoprostol, partially reproduced this phenotype. Together these data led us to hypothesize that immune-derived prostaglandin E₂ raised from the insect response to the fungal challenge is involved in disturbing follicle development, contributing to a reduction in host reproductive output and acting as a host-derived adaptive effector to infection.     

Functional properties of type I and type II cytochromes c3 from Desulfovibrio africanus

Type I cytochrome c₃ is a key protein in the bioenergetic metabolism of Desulfovibrio spp., mediating electron transfer between periplasmic hydrogenase and multihaem cytochromes associated with membrane bound complexes, such as type II cytochrome c₃. This work presents the NMR assignment of the haem substituents in type I cytochrome c₃ isolated from Desulfovibrio africanus and the thermodynamic and kinetic characterisation of type I and type II cytochromes c₃ belonging to the same organism. It is shown that the redox properties of the two proteins allow electrons to be transferred between them in the physiologically relevant direction with the release of energised protons close to the membrane where they can be used by the ATP synthase.     

In silico and in vitro characterization of phospholipase A2 isoforms from soybean (Glycine max)

At the present, no secreted phospholipase A₂ (sPLA₂) from soybean (Glycine max) was investigated in detail. In this work we identified five sequences of putative secreted sPLA₂ from soybean after a BLAST search in G. max database. Sequence analysis showed a conserved PA2c domain bearing the Ca²⁺ binding loop and the active site motif. All the five mature proteins contain 12 cysteine residues, which are commonly conserved in plant sPLA₂s. We propose a phylogenetic tree based on sequence alignment of reported plant sPLA₂s including the novel enzymes from G. max. According to PLA₂ superfamily, two of G. max sPLA₂s are grouped as XIA and the rest of sequences as XIB, on the basis of differences found in their molecular weights and deviating sequences especially in the N- and C-terminal regions of the isoenzymes. Furthermore, we report the cloning, expression and purification of one of the putative isoenzyme denoted as GmsPLA₂-XIA-1. We demonstrate that this mature sPLA₂ of 114 residues had PLA₂ activity on Triton:phospholipid mixed micelles and determine the kinetic parameters for this system. We generate a model based on the known crystal structure of sPLA₂ from rice (isoform II), giving first insights into the three-dimensional structure of folded GmsPLA₂-XIA-1. Besides describing the spatial arrangement of highly conserved pair HIS-49/ASP-50 and the Ca⁺² loop domains, we propose the putative amino acids involved in the interfacial recognition surface. Additionally, molecular dynamics simulations indicate that calcium ion, besides its key function in the catalytic cycle, plays an important role in the overall stability of GmsPLA₂-XIA-1 structure.     

Tuning of functional heme reduction potentials in Shewanella fumarate reductases

The fumarate reductases from S. frigidimarina NCIMB400 and S. oneidensis MR-1 are soluble and monomeric enzymes located in the periplasm of these bacteria. These proteins display two redox active domains, one containing four c-type hemes and another containing FAD at the catalytic site. This arrangement of single-electron redox co-factors leading to multiple-electron active sites is widespread in respiratory enzymes. To investigate the properties that allow a chain of single-electron co-factors to sustain the activity of a multi-electron catalytic site, redox titrations followed by NMR and visible spectroscopies were applied to determine the microscopic thermodynamic parameters of the hemes. The results show that the redox behaviour of these fumarate reductases is similar and dominated by a strong interaction between hemes II and III. This interaction facilitates a sequential transfer of two electrons from the heme domain to FAD via heme IV.     

Electron transfer patterns of the di-heme protein cytochrome c4 from Pseudomonas stutzeri

We report kinetic data for the two-step electron transfer (ET) oxidation and reduction of the two-domain di-heme redox protein Pseudomonas stutzeri cytochrome (cyt) c₄ by [Co(bipy)₃]^2+/3+ (bipy = 2,2′-bipyridine). Following earlier reports, the data accord with both bi- and tri-exponential kinetics. A complete kinetic scheme includes both “cooperative” intermolecular ET between each heme group and the external reaction partner, and intramolecular ET between the two heme groups. A new data analysis scheme shows unequivocally that two-ET oxidation and reduction of P. stutzeri cyt c₄ is entirely dominated by intermolecular ET between the heme groups and the external reaction partner in the ms time range, with virtually no contribution from intramolecular interheme ET in this time range. This is in striking contrast to two-ET electrochemical oxidation or reduction of P. stutzeri cyt c₄ for which fast, ms to sub-ms intramolecular interheme ET is a crucial step. The rate constant dependence on the solvent viscosity has disclosed strong coupling to both a (set of) frictionally damped solvent/protein nuclear modes and intramolecular friction-less “ballistic” modes, indicative of notable protein structural mobility in the overall two-ET process. We suggest that conformational protein mobility blocks intramolecular interheme ET in bulk homogeneous solution but triggers opening of this gated ET channel in the electrochemical environment or in the membrane environment of natural respiratory cyt c₄ function.     

Interactions between TGF-β1 and TGF-β3 and their role in medial edge epithelium cell death and palatal fusion in vitro

In recent decades, studies have shown that both TGF-β₁ and TGF-β₃ play an important role in the induction of medial edge epithelium (MEE) cell death and palatal fusion. Many of these experiments involved the addition or blockage of one of these growth factors in wild-type (WT) mouse palate cultures, where both TGF-β₁ and TGF-β₃ are present. Few studies have addressed the existence of interactions between TGF-β₁ and TGF-β₃, which could modify their individual roles in MEE cell death during palatal fusion. We carried out several experiments to test this possibility, and to investigate how this could influence TGF-β₁ and TGF-β₃ actions on MEE cell death and palatal shelf fusion. We double-immunolabelled developing mouse palates with anti-TGF-β₁ or anti-TGF-β₃ antibodies and TUNEL, added rhTGF-β₁ or rhTGF-β₃ or blocked the TGF-β₁ and TGF-β₃ action at different concentrations to WT or Tgf-β₃ null mutant palate cultures, performed in situ hybridizations with Tgf-β₁ or Tgf-β₃ riboprobes, and measured the presence of TUNEL-positive midline epithelial seam (MES) cells and MES disappearance (palatal shelf fusion) in the different in vitro conditions. By combining all these experiments, we demonstrate great interaction between TGF-β₁ and TGF-β₃ in the developing palate and confirm that TGF-β₃ has a more active role in MES cell death than TGF-β₁, although both are major inductors of MES disappearance. Finally, the co-localization of TGF-β₁, but not TGF-β₃, with TUNEL in the MES allows us to suggest a possible role for TGF-β₁ in MES apoptotic clearance.     

Parameters determining the relative efficacy of hydroxy-naphthoquinone inhibitors of the cytochrome bc1 complex

Hydroxy-naphthoquinones are competitive inhibitors of the cytochrome bc₁ complex that bind to the ubiquinol oxidation site between cytochrome b and the iron-sulfur protein and presumably mimic a transition state in the ubiquinol oxidation reaction catalyzed by the enzyme. The parameters that affect efficacy of binding of these inhibitors to the bc₁ complex are not well understood. Atovaquone®, a hydroxy-naphthoquinone, has been used therapeutically to treat Pneumocystis carinii and Plasmodium infections. As the pathogens have developed resistance to this drug, it is important to understand the molecular basis of the drug resistance and to develop new drugs that can circumvent the drug resistance. We previously developed the yeast and bovine bc₁ complexes as surrogates to model the interaction of atovaquone with the bc₁ complexes of the target pathogens and human host. As a first step to identify new cytochrome bc₁ complex inhibitors with therapeutic potential and to better understand the determinants of inhibitor binding, we have screened a library of 2-hydroxy-naphthoquinones with aromatic, cyclic, and non-cyclic alkyl side-chain substitutions at carbon-3 on the hydroxy-quinone ring. We found a group of compounds with alkyl side-chains that effectively inhibit the yeast bc₁ complex. Molecular modeling of these into the crystal structure of the yeast cytochrome bc₁ complex provides structural and quantitative explanations for their binding efficacy to the target enzyme. In addition we also identified a 2-hydroxy-naphthoquinone with a branched side-chain that has potential for development as an anti-fungal and anti-parasitic therapeutic.     

Quercetin-induced H2O2 mediates the pathogen resistance against Pseudomonas syringae pv. Tomato DC3000 in Arabidopsis thaliana

Quercetin is a potent antioxidant and has been extensively used as a therapy intervention to prevent age-associated diseases. However, emerging studies showed it can also act as a prooxidant and induce H₂O₂ under certain conditions. In the current study, our results showed that quercetin contributed to the pathogen resistance in Arabidopsis thaliana (Arabidopsis) in response to the infection of virulent strain Pseudomonas syringae pv. Tomato DC3000 (Pst). Various defense responses, such as H₂O₂ burst, callose deposition, cell death, PR1 (pathogenesis-related 1) and PAL1 (Phe ammonia-lyase 1) gene expression, have been investigated in quercetin-pretreated Pst-inoculated Arabidopsis Col-0 and there was a strong defensive response in quercetin-pretreated Arabidopsis against virulent Pst. However, with the presence of catalase, the protective effects of quercetin on pathogen resistance to virulent Pst disappeared in Arabidopsis, suggesting that H₂O₂ may play a key role in plant defense responses. In addition, we confirmed that quercetin did not show any beneficial effect on pathogen-free leaves in Arabidopsis, indicating that pathogen challenge is also required to induce the defense responses in quercetin-pretreated Arabidopsis. Furthermore, strong defense responses have been observed in quercetin-pretreated Arabidopsis mutant jar1, ein2, and abi1-2 under Pst challenge, whereas no protective effect has been observed in quercetin-pretreated Arabidopsis mutant NahG and npr1. These findings indicate that quercetin induces the resistance to Pst in Arabidopsis via H₂O₂ burst and involvement of SA and NPR1.