Cross-talks between Ca<Superscript>2+</Superscript>/CaM and H<Subscript>2</Subscript>O<Subscript>2</Subscript> in abscisic acid-induced antioxidant defense in leaves of maize plants exposed to water stress

Here we examined whether Ca²⁺/Calmodulin (CaM) is involved in abscisic acid (ABA)-induced antioxidant defense and the possible relationship between CaM and H₂O₂ in ABA signaling in leaves of maize (Zea mays L.) plants exposed to water stress. An ABA-deficient mutant vp5 and its wild type were used for the experimentation. We found that water stress enhanced significantly the contents of CaM and H₂O₂, and the activities of chloroplastic and cytosolic superoxide dismutase (SOD), ascorbate peroxidase (APX) and glutathione reductase (GR), and the gene expressions of the CaM1, cAPX, GR1 and SOD4 in leaves of wild-type maize. However, the increases mentioned above were almost arrested in vp5 plants and in the wild-type plants pretreated with ABA biosynthesis inhibitor tungstate (T), suggesting that ABA is required for water stress-induced H₂O₂ production, the enhancement of CaM content and antioxidant defense. Besides, we showed that the up-regulation of water stress-induced antioxidant defense was almost completely blocked by pretreatment with Ca²⁺ inhibitors, CaM antagonists and reactive oxygen (ROS) manipulators. Moreover, the analysis of time course of CaM and H₂O₂ production under water stress showed that the increase in CaM content preceded that of H₂O₂. These results suggested that Ca²⁺/CaM and H₂O₂ were involved in the ABA-induced antioxidant defense under water stress, and the increases of Ca²⁺/CaM contents triggered H₂O₂ production, which inversely affected the contents of CaM. Thus, a cross-talk between Ca²⁺/CaM and H₂O₂ may play a pivotal role in the ABA signaling.     

Abscisic acid-induced apoplastic H<Subscript>2</Subscript>O<Subscript>2</Subscript> accumulation up-regulates the activities of chloroplastic and cytosolic antioxidant enzymes in maize leaves

The histochemical and cytochemical localization of abscisic acid (ABA)-induced H₂O₂ production in leaves of maize (Zea mays L.) plants were examined, using 3,3-diaminobenzidine (DAB) and CeCl₃ staining, respectively, and the relationship between ABA-induced H₂O₂ production and ABA-induced subcellular activities of antioxidant enzymes was studied. H₂O₂ generated in response to ABA treatment was detected within 0.5 h in major veins of the leaves and maximized at about 2–4 h. In mesophyll and bundle sheath cells, ABA-induced H₂O₂ accumulation was observed only in apoplast, and the greatest accumulation occurred in the walls of mesophyll cells facing large intercellular spaces. Meanwhile, ABA treatment led to a significant increase in the activities of the leaf chloroplastic and cytosolic antioxidant enzymes superoxide dismutase (SOD), ascorbate peroxidase (APX) and glutathione reductase (GR), and pretreatment with the NADPH oxidase inhibitor diphenyleneiodonium (DPI), the O ₂ ⁻ scavenger Tiron and the H₂O₂ scavenger dimethylthiourea (DMTU) almost completely arrested the increase in the activities of these antioxidant enzymes. Our results indicate that the accumulation of apoplastic H₂O₂ is involved in the induction of the chloroplastic and cytosolic antioxidant enzymes. Moreover, an oxidative stress induced by paraquat (PQ), which generates O ₂ ⁻ and then H₂O₂ in chloroplasts, also up-regulated the activities of the chloroplastic and cytosolic antioxidant enzymes, and the up-regulation was blocked by the pretreatment with Tiron and DMTU. These data suggest that H₂O₂ produced at a specific cellular site could coordinate the activities of antioxidant enzymes in different subcellular compartments.     

外源亚精胺对Ca(NO3)2胁迫下番茄幼苗光合特性和抗氧化酶活性的影响

以耐盐性较弱的番茄品种上海‘合作903’幼苗为试验材料,采用营养液栽培方法,研究了叶面喷施1mmol.L-1亚精胺(Spd)对75mmol.L-1 Ca(NO32)胁迫下番茄幼苗生长、叶绿素含量、光合及荧光参数、叶片抗氧化酶活性的影响,以探讨外源亚精胺缓解Ca(NO3)2胁迫伤害的机制。结果显示:Ca(NO3)2胁迫能够显著抑制番茄幼苗的生长;与Ca(NO32)胁迫处理相比,叶面喷施外源Spd 9d后,受胁迫番茄幼苗的株高、茎粗、干重、鲜重分别显著增加70.9%、15.8%、43.4%、41.4%;叶绿素a、b的含量分别提高17.7%、13.8%;净光合速率(Pn)、气孔导度(Gs)、蒸腾速率(Tr)、光合电子传递效率(rETR)和光化学猝灭系数(qP)分别升高6.6%、18.0%、31.0%、4.9%、5.0%,而胞间二氧化碳浓度(Ci)、非光化学猝灭系数(qN)分别降低21.5%、8.1%;超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性分别增加10.7%和37.5%,而丙二醛(MDA)含量和超氧阴离子(O2.-)产生速率分别显著下降34.5%、17.1%。研究表明,外源Spd通过提高番茄幼苗抗氧化酶活性来有效清除体内活性氧,维持光合机构的稳定性,提高其光合速率,从而缓解Ca(NO3)2胁迫对番茄幼苗的伤害。     

等渗NaCl和Ca(NO_3)_2胁迫对黄瓜幼苗生长和生理特性的影响

以盐敏感型的黄瓜品种‘津春2号’为材料,采用营养液栽培方法,研究了等渗NaCl和Ca(NO3)2胁迫对黄瓜幼苗生长和生理特性的影响。结果表明,NaCl胁迫不仅抑制黄瓜幼苗地上部生长,而且对根系生长也具有抑制作用;而Ca(NO3)2胁迫主要抑制黄瓜幼苗地上部生长,对根系生长没有明显影响。NaCl胁迫下,黄瓜幼苗净光合速率(Pn)降低不仅由非气孔因素引起,而且碳同化能力也降低;而Ca(NO3)2胁迫下Pn降低主要由非气孔因素引起,对植株的碳同化能力没有明显影响。等渗NaCl和Ca(NO3)2胁迫导致黄瓜幼苗叶片光系统Ⅱ(PSⅡ)反应中心过剩光能(Ex)显著升高,致使叶片超氧阴离子自由基(O-·2)产生速率加快,过氧化氢(H2O2)含量显著升高,但NaCl胁迫的升高幅度均大于Ca(NO3)2胁迫。NaCl胁迫在提高过氧化物酶(POD)和过氧化氢酶(CAT)活性的同时,降低了超氧化物歧化酶(SOD)活性,对活性氧(ROS)的清除能力有限,致使叶片ROS大量积累,膜质过氧化伤害严重;而Ca(NO3)2胁迫显著提高了SOD、POD和CAT活性,有效清除了ROS,对叶片未造成严重的膜质过氧化伤害。NaCl和Ca(NO3)2胁迫分别导致黄瓜幼苗体内Na+和Ca2+大量积累,致使离子平衡被打破,但过多Na+要比过多的Ca2+对植物体的伤害作用大。上述结果说明,黄瓜幼苗对等渗NaCl和Ca(NO3)2胁迫的生理响应存在差异。NaCl胁迫下黄瓜幼苗体内Na+大量积累,不仅限制了环境中CO2进入叶肉细胞,而且使碳同化能力降低,加之ROS清除能力有限,不能有效清除PSⅡ反应中心过剩能量所导致的ROS,因而产生了严重的膜质过氧化,影响了植株生长和光合速率;而Ca(NO3)2胁迫下,黄瓜幼苗碳同化能力没有受到影响,ROS也能够被抗氧化系统有效清除,植株没有产生严重的膜质过氧化,因而对生长和光合速率的影响没有NaCl胁迫大。     

不同BR施用方式诱导黄瓜幼苗对Ca(NO_3)_2胁迫抗性的研究

为探讨外源油菜素内酯(brassinosteroid,BR)诱导黄瓜幼苗对Ca(NO3)2胁迫抗性的效果,研究了3种外源BR施用方法(0.01mg·L-1 BR浸种、0.1mg·L-1 BR喷叶及其二者结合施用)对Ca(NO3)2胁迫(60mmol·L-1)下黄瓜幼苗生长、生理活动以及光合作用的影响。结果表明:(1)3种外源BR方法处理后,Ca(NO3)2胁迫下的黄瓜幼苗株高、茎粗、展开叶片数、叶面积、干重含水量均显著提高,同时其叶片游离脯氨酸和可溶性糖含量上升,过氧化物酶活性提高,而其丙二醛(MDA)含量趋于无Ca(NO3)2胁迫对照的水平;(2)外源BR处理还提高了Ca(NO3)2胁迫下黄瓜幼苗的净光合速率、蒸腾速率和气孔导度,却抑制了Ca(NO3)2胁迫下胞间CO2浓度的升高。研究认为,适宜浓度的外源BR浸种和喷叶处理均可有效增强黄瓜幼苗渗透调节能力,降低细胞膜质过氧化伤害程度,提高抗氧化酶活性和光合效率,从而表现出对Ca(NO3)2胁迫的抗性,并以操作简便、用量极低的0.01mg·L-1 BR浸种方法效果最佳。     

P2X<Subscript>2</Subscript> and P2X<Subscript>3</Subscript> purinoceptors in the rat enteric nervous system

Adenosine 5-triphosphate receptors are known to be involved in fast excitatory postsynaptic currents in myenteric neurons of the digestive tract. In the present study, the distribution of P2X₂ and P2X₃ receptor mRNA was examined by in situ hybridisation while P2X₂ and P2X₃ receptor protein was localised by immunohistochemical methods. In addition, P2X₂ and P2X₃ receptors were colocalised with calbindin and calretinin in the myenteric and submucosal plexus. P2X₂- and P2X₃-immunoreactive neurons were found in the myenteric and submucosal plexuses throughout the entire length of the rat digestive tract from the stomach to the colon. Approximately 60%, 70% and 50% of the ganglion cells in the myenteric plexus of the gastric corpus, ileum and distal colon, and 56% and 45% in the submucosal plexus of the ileum and distal colon, respectively, showed positive immunoreactivity to the P2X₂ receptor. Approximately 10%, 2% and 15% of the ganglion cells in the myenteric plexus of the gastric corpus, ileum and distal colon, and 62% and 40% in the submucosal plexus of the ileum and distal colon, respectively, showed positive immunoreactivity to the P2X₃ receptor. Double-labelling studies showed that about 10–25% of the neurons with P2X₂ immunoreactivity in myenteric plexus and 30–50% in the submucosal plexus were found to express calbindin or calretinin. About 80% of the neurons with P2X₃ receptor immunoreactivity in the myenteric plexus and about 40% in the submucosal plexus expressed calretinin. Approximately 30–75% of the neurons with P2X₃ receptor immunoreactivity in the submucosal plexus expressed calbindin, while none of them were found to express calbindin in the myenteric plexus.     

Neutrophil Gelatinase-Associated Lipocalin induces the expression of heme oxygenase-1 and superoxide dismutase <Subscript>1, 2</Subscript>

Lipocalin-2 (Lcn2, NGAL) is a member of the lipocalin super family with diverse function such as the induction of apoptosis, the suppression of bacterial growth, and modulation of inflammatory response. Much interest has recently been focused on the physiological/pathological role of the lipocalin-2 that is considered to be a novel protective factor against oxidative stress. However, its precise biological roles in this protection are not fully understood. In this report we intended to test the effect of lipocalin-2 on the expression of heme oxygenase _{(1, 2)} and superoxide dismutase _{(1, 2)} which are two strong antioxidants. NGAL was cloned to pcDNA3.1 plasmid by using genetic engineering method. The recombinant vector was transfected to CHO and HEK293T to establish stable cell expressing NGAL and the expression of HO-1, 2 and SOD_{1, 2} were compared with appropriate controls by RT-PCR and western blot. On the other hand, expression of NGAL was suppressed by siRNA transfection in order to study the effect of lipocalin-2 on mentioned genes/proteins. The results showed that the expression of HO-1 and SOD_{1, 2} enzymes were higher in cells expressing recombinant lipocalin-2 compared with the control cells. Although the expression of HO-1 was lower in NGAL silencing cells, the expression of SOD₁ and SOD₂ were higher. Our data suggest that NGAL is a potent inducer of HO-1 and somewhat SOD₁ and SOD₂ and it appears that part of antioxidant property of NGAL could be attributed to the induction of HO-1and SOD_{1, 2}.     

Studies on DNA binding to metal complexes of Sal<Subscript>2</Subscript>trien

The complexes [Fe(Sal2trien)]NO3 and Cu(Sal2trien) have been synthesized and their interaction with calf thymus DNA has been investigated for the first time using UV spectra, fluorescence spectra, thermal denaturation, and viscosity measurements. The experimental results show conformably that the mode of binding of the complex [Fe(Sal2trien)]NO3 to DNA is nonclassical electrostatic action, but the mode of binding of the complex Cu(Sal2trien) to DNA is classical intercalation.     

Effects of elevated CO<Subscript>2</Subscript> and/or O<Subscript>3</Subscript> on hormone IAA in needles of Chinese pine

This study examined the effects of season-long exposure of Chinese pine (Pinus tabulaeformis) to elevated carbon dioxide (CO₂) and/or ozone (O₃) on indole-3-acetic acid (IAA) content, activities of IAA oxidase (IAAO) and peroxidase (POD) in needles. Trees grown in open-top chambers (OTC) were exposed to control (ambient O₃, 55 nmol mol⁻¹ + ambient CO₂, 350 μmol mol⁻¹, CK), elevated CO₂ (ambient O₃ + high CO₂, 700 μmol mol⁻¹, EC) and elevated O₃ (high O₃, 80 ± 8 nmol mol⁻¹ + ambient CO₂, EO) OTCs from 1 June to 30 September. Plants grown in elevated CO₂ OTC had a growth increase of axial shoot and needle length, compared to control, by 20% and 10% respectively, while the growth in elevated O₃ OTC was 43% and 7% less respectively, than control. An increase in IAA content and POD activity and decrease in IAAO activity were observed in trees exposed to elevated CO₂ concentration compared with control. Elevated O₃ decreased IAA content and had no significant effect on IAAO activity, but significantly increased POD activity. When trees pre-exposed to elevated CO₂ were transferred to elevated O₃ (EC–EO) or trees pre-exposed to elevated O₃ were transferred to elevated CO₂ (EO–EC), IAA content was lower while IAAO activity was higher than that transferred to CK (EC–CK or EO–CK), the change in IAA content was also related to IAAO activity. The results indicated that IAAO and POD activities in Chinese pine needles may be affected by the changes in the atmospheric environment, resulting in the change of IAA metabolism which in turn may cause changes in Chinese pine’s growth. An erratum to this article can be found at     

Cloning a Truncated Fragment (stx2a<Subscript>1</Subscript>) of the Shiga-Like Toxin 2A<Subscript>1</Subscript> Subunit of EHEC O157:H7: Candidate Immunogen for a Subunit Vaccine

Shiga toxins consist of enzymatically active A and B subunit multimers. The A subunit of shiga-like toxins can be proteolytically cleaved into two parts, A₁ and A₂, with A₁ being responsible for toxic activity. Antibody neutralizing the A₁ subunit of shiga toxin may protect against infection of the enterohemorrhagic Escherichia coli (EHEC O157:H7). It was difficult to express the full-length A₁ subunit of shiga toxin 2 (stx2A₁) in a previous study. We have now analyzed the full-length of stx2A₁ using bioinformatics software. The data show that the carboxyl terminal (of ~15 amino-acid residues) has strong hydrophobicity and low antigenicity. We cloned and expressed a truncated fragment of stx2A₁ (15 amino-acid residues of the carboxyl terminal being removed), designated stx2a₁, which can evoke a humoral immune response. Anti-Stx2a₁ antibodies can neutralize the native shiga toxin 2 both in vivo and in vitro, which suggests that Stx2a₁ serves as a candidate immunogen for a subunit vaccine that can also be used as the antigen to screen phage anti-shiga toxin antibody libraries. L. Liu and H. Zeng contributed equally to this study.     

Changes in antioxidant status of myocardium during oxidative stress under the influence of coenzyme Q<Subscript>10</Subscript>

Changes in myocardium were studied during oxidative stress induced by infusion of hydrogen peroxide in the coronary vessels of isolated rat heart. Moderate concentrations of H2O2 increased the heart rate but decreased the contractile force, whereas higher concentrations of H2O2 decreased both parameters and increased the end diastolic pressure. The effect of H2O2 was stable, cumulative, and was associated with disturbance in respiration of mitochondria, increased production of ROS in them, and decrease in activities of antioxidant enzymes in the myocardium. Changes in the antioxidant status of the myocardium induced by long-term addition of coenzyme Q(10) into food was accompanied by decrease in the negative inotropic effect of H2O2, whereas the levels of superoxide dismutase and glutathione peroxidase after oxidative stress were virtually unchanged. The activities of these enzymes displayed a high positive correlation with the cardiac function. The findings suggest that coenzyme Q(10) should increase resistance of the myocardium to oxidative stress not only by a direct antioxidant mechanism but also indirectly, due to increased protection of antioxidant enzymes.     

The effect of cadmium on CO<Subscript>2</Subscript> exchange,variable fluorescence of chlorophyll,and the level of antioxidant enzymes in pea leaves

CO₂ exchange, variable chlorophyll fluorescence, the intensity of lipid peroxidation (POL), and the activity of antioxidant enzymes in leaves of two-week-old pea seedlings (Pisum sativum L.) exposed to 0.01, 0.1, and 1 mM aqueous solutions of Cd(NO₃)₂ for 2 h were studied. Incubation with Cd²⁺ ions resulted in a reduction of the maximum quantum efficiency of photosynthesis, CO₂ uptake rate, and photosystem II (PSII) activity, as assessed by F _v/F ₀ ratio. The intensity of POL in leaves of all treated seedlings was below or close to the control level. The activity of superoxide dismutase (SOD) and glutathione reductase (GR) increased in all treatments; the activity of ascorbate peroxidase (AP) exceeded the control level only in seedlings exposed to the high Cd²⁺ concentration (1 mM), and the activity of peroxidase increased at the low concentration (0.01 mM). We found that the reduction in the activity of the photosynthetic apparatus under conditions of 2-h-long Cd²⁺-induced stress was not related to an intensification of POL processes. It was concluded that stimulation of the activity of antioxidant enzymes—SOD, GR, AP, and peroxidase—is a pathway for pea plant adaptation to toxic effect of cadmium.Translated from Fiziologiya Rastenii, Vol. 52, No. 1, 2005, pp. 21–26.Original Russian Text Copyright © 2005 by Balakhnina, Kosobryukhov, Ivanov, Kreslavskii.     

P2X<Subscript>3</Subscript> Receptor Involvement in Pain States

The understanding of how pain is processed at each stage in the peripheral and central nervous system is the precondition to develop new therapies for the selective treatment of pain. In the periphery, ATP can be released from various cells as a consequence of tissue injury or visceral distension and may stimulate the local nociceptors. The highly selective distribution of P2X₃ and P2X_2/3 receptors within the nociceptive system has inspired a variety of approaches to elucidate the potential role of ATP as a pain mediator. Depolarization by ATP of neurons in pain–relevant neuronal structures such as trigeminal ganglion, dorsal root ganglion, and spinal cord dorsal horn neurons are well investigated. P2X receptor-mediated afferent activation appears to have been implicated in visceral and neuropathic pain and even in migraine and cancer pain. This article reviews recently published research describing the role that ATP and P2X receptors may play in pain perception, highlighting the importance of the P2X₃ receptor in different states of pain.     

In<Emphasis Type="Italic"> Saccharomyces cerevisiae</Emphasis>, the effect of H<Subscript>2</Subscript>O<Subscript>2</Subscript> on ATP,but not on glyceraldehyde-3-phosphate dehydrogenase,depends on the glucose concentration

As has been previously shown, Saccharomyces cerevisiae grown in 2% or 0.025% glucose uses this carbohydrate by the fermentative or oxidative pathways, respectively. Depending on the glucose concentration in the medium, the effect of the addition of H₂O₂ on the level of ATP and on glyceraldehyde-3-phosphate dehydrogenase (GAPDH) activity differed. In the presence of 2% glucose, ATP and GAPDH decreased sharply during the first few minutes of treatment, whereas in the presence of 0.025% glucose, GAPDH activity decreased similarly, but the ATP level remained practically unchanged. The addition of 3 mM glutathione to the culture media prevented the depletion of ATP levels and GAPDH activity in the presence of H₂O₂. Catalase and superoxide dismutase activities did not vary significantly when yeast cells were grown either in 2% or in 0.025% glucose.     

Reduction of Fe(II)EDTA-NO by a newly isolated <Emphasis Type="Italic">Pseudomonas</Emphasis> sp. strain DN-2 in NO<Subscript><Emphasis Type="Italic">x</Emphasis></Subscript> scrubber solution

Biological reduction of nitric oxide (NO) chelated by ferrous ethylenediaminetetraacetate (Fe(II)EDTA) to N₂ is one of the core processes in a chemical absorption–biological reduction integrated technique for nitrogen oxide (NO _x ) removal from flue gases. A new isolate, identified as Pseudomonas sp. DN-2 by 16S rRNA sequence analysis, was able to reduce Fe(II)EDTA-NO. The specific reduction capacity as measured by NO was up to 4.17 mmol g DCW⁻¹ h⁻¹. Strain DN-2 can simultaneously use glucose and Fe(II)EDTA as electron donors for Fe(II)EDTA-NO reduction. Fe(III)EDTA, the oxidation of Fe(II)EDTA by oxygen, can also serve as electron acceptor by strain DN-2. The interdependency between various chemical species, e.g., Fe(II)EDTA-NO, Fe(II)EDTA, or Fe (III)EDTA, was investigated. Though each complex, e.g., Fe(II)EDTA-NO or Fe(III)EDTA, can be reduced by its own dedicated bacterial strain, strain DN-2 capable of reducing Fe(III)EDTA can enhance the regeneration of Fe(II)EDTA, hence can enlarge NO elimination capacity. Additionally, the inhibition of Fe(II)EDTA-NO on the Fe(III)EDTA reduction has been explored previously. Strain DN-2 is probably one of the major contributors for the continual removal of NO _x due to the high Fe(II)EDTA-NO reduction rate and the ability of Fe(III)EDTA reduction.     

Long-term effects of elevated atmospheric CO<Subscript>2</Subscript> on species composition and productivity of a southern African C<Subscript>4</Subscript> dominated grassland in the vicinity of a CO<Subscript>2</Subscript> exhalation

We describe the long-term effects of a CO₂ exhalation, created more than 70 years ago, on a natural C₄ dominated sub-tropical grassland in terms of ecosystem structure and functioning. We tested whether long-term CO₂ enrichment changes the competitive balance between plants with C₃ and C₄ photosynthetic pathways and how CO₂ enrichment has affected species composition, plant growth responses, leaf properties and soil nutrient, carbon and water dynamics. Long-term effects of elevated CO₂ on plant community composition and system processes in this sub-tropical grassland indicate very subtle changes in ecosystem functioning and no changes in species composition and dominance which could be ascribed to elevated CO₂ alone. Species compositional data and soil δ¹³C isotopic evidence suggest no detectable effect of CO₂ enrichment on C₃:C₄ plant mixtures and individual species dominance. Contrary to many general predictions C₃ grasses did not become more abundant and C₃ shrubs and trees did not invade the site. No season length stimulation of plant growth was found even after 5 years of exposure to CO₂ concentrations averaging 610 μmol mol⁻¹. Leaf properties such as total N decreased in the C₃ but not C₄ grass under elevated CO₂ while total non-structural carbohydrate accumulation was not affected. Elevated CO₂ possibly lead to increased end-of-season soil water contents and this result agrees with earlier studies despite the topographic water gradient being a confounding problem at our research site. Long-term CO₂ enrichment also had little effect on soil carbon storage with no detectable changes in soil organic matter found. There were indications that potential soil respiration and N mineralization rates could be higher in soils close to the CO₂ source. The conservative response of this grassland suggests that many of the reported effects of elevated CO₂ on similar ecosystems could be short duration experimental artefacts that disappear under long-term elevated CO₂ conditions.     

Cellular expression of C<Subscript>3</Subscript> and C<Subscript>4</Subscript> photosynthetic enzymes in the amphibious sedge<Emphasis Type="Italic"> Eleocharis retroflexa</Emphasis> ssp.<Emphasis Type="Italic"> chaetaria</Emphasis>

The amphibious leafless sedge Eleocharis retroflexa ssp. chaetaria expresses C₄-like biochemical characteristics in both the terrestrial and submerged forms. Culms of the terrestrial form have Kranz anatomy, whereas those of the submerged form have Kranz-like anatomy combined with anatomical features of aquatic plant leaves. We examined the immunolocalization of C₃ and C₄ enzymes in culms of the two forms. In both forms, phosphoenolpyruvate carboxylase; pyruvate, Pi dikinase; and NAD-malic enzyme were compartmentalized between the mesophyll (M) and Kranz cells, but their levels were somewhat reduced in the submerged form. In the terrestrial form, ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco) occurred mainly in the Kranz cells, and weakly in the M chloroplasts. In the submerged form, the rubisco occurred at higher levels in the M cells than in the terrestrial form. In both forms, the C₄ pattern of enzyme expression was clearer in the M cells adjacent to Kranz cells than in distant M cells. During the transition from terrestrial to submerged conditions, the enzyme expression pattern changed in submerged mature culms that had been formed in air before submergence, and matched that in culms newly developed underwater. It seems that effects of both environmental and developmental factors overlap in the C₄ pattern expression in this plant.     

Inhibitory role of TGIF in the As<Subscript>2</Subscript>O<Subscript>3</Subscript>-regulated p21<Superscript><Emphasis Type="Italic">WAF1/CIP1</Emphasis></Superscript> expression

Although arsenic is an infamous carcinogen, it has been effectively used to treat acute promyelocytic leukemia, and can induce cell cycle arrest or apoptosis in human solid tumors. Previously, we had demonstrated that opposing effects of ERK1/2 and JNK on p21 expression in response to arsenic trioxide (As₂O₃) are mediated through the Sp1 responsive elements of the p21 promoter in A431 cells. Presently, we demonstrate that Sp1, and c-Jun functionally cooperate to activate p21 promoter expression through Sp1 binding sites (−84/−64) by using DNA affinity binding, chromatin immunoprecipitation, and promoter assays. Surprisingly, As₂O₃-induced c-Jun(Ser63/73) phosphorylation can recruit TGIF/HDAC1 to the Sp1 binding sites and then suppress p21 promoter activation. We suggest that, after As₂O₃ treatment, the N-terminal domain of c-Jun phosphorylation by JNK recruits TGIF/HDAC1 to the Sp1 sites and then represses p21 expression. That is, TGIF is involved in As₂O₃-inhibited p21 expression, and then blocks the cell cycle arrest.     

外源一氧化氮对硝酸盐胁迫下黄瓜幼苗生长及抗氧化酶活性的影响

采用营养液培养试验,通过添加不同浓度(0.05、0.1、0.2、0. mmol·L^-1)的硝普钠（SNP）作为NO供体,研究外源NO对NO₃-胁迫下黄瓜幼苗生长及叶片抗氧化酶活性的影响.结果表明： 140 mmol·L^-1 NO₃-胁迫下,外加0.1 mmol·L^-1 SNP处理1 d和7 d后,黄瓜幼苗叶片中SOD、CAT和APX活性显著升高;MDA含量显著降低,可溶性蛋白含量增加,说明外加0.1 mmol·L^-1 SNP增强了黄瓜幼苗对活性氧的清除能力,降低了膜脂过氧化程度,幼苗生长势增加,对高浓度NO₃-胁迫的抗性增强;当SNP浓度达0.3 mmol·L^-1处理7 d后,SOD、POD、CAT和APX活性均开始降低,MDA含量增加,黄瓜幼苗受害加重.外加一定浓度(0.1～0.2 mmol·L^-1)的外源NO可缓解NO₃^-胁迫对黄瓜幼苗的影响.     

Variations of vinblastine accumulation and redox state affected by exogenous H<Subscript>2</Subscript>O<Subscript>2</Subscript> in <Emphasis Type="Italic">Catharanthus roseus</Emphasis> (L.) G. Don

Effects of exogenous H₂O₂ application on vinblastine (VBL) and its precursors, vindoline (VIN), catharanthine (CAT) and α-3′,4′-anhydrovinblastine (AVBL), were measured in Catharanthus roseus seedlings in order to explore possible correlation of VBL formation with oxidative stress. VBL accumulation has previously been shown to be regulated by an in vitro H₂O₂-dependent peroxidase (POD)-like synthase. Experimental exposure of plants to different concentrations of H₂O₂ showed that endogenous H₂O₂ and alkaloid concentrations in leaves were positively elevated. The time-course variations of alkaloid concentrations and redox state, reflected by the concentrations of H₂O₂, ascorbic acid (AA), oxidative product of glutathione (GSSG) and POD activity, were significantly altered due to H₂O₂ application. The further correlation analysis between alkaloids and redox status indicated that VBL production was tightly correlated with redox status. These results provide a new link between VBL metabolisms and redox state in C. roseus.