Foliar application of ascorbic acid enhances salinity stress tolerance in barley (Hordeum vulgare L.) through modulation of morpho-physio-biochemical attributes,ions uptake,osmo-protectants and stress response genes expression

Barley (Hordeum vulgare L.) is a major cereal grain and is known as a halophyte (a halophyte is a salt-tolerant plant that grows in soil or waters of high salinity). We therefore conducted a pot experiment to explore plant growth and biomass, photosynthetic pigments, gas exchange attributes, stomatal properties, oxidative stress and antioxidant response and their associated gene expression and absorption of ions in H. Vulgare. The soil used for this analysis was artificially spiked at different salinity concentrations (0, 50, 100 and 150 mM) and different levels of ascorbic acid (AsA) were supplied to plants (0, 30 and 60 mM) shortly after germination of the seed. The results of the present study showed that plant growth and biomass, photosynthetic pigments, gas exchange parameters, stomatal properties and ion uptake were significantly (p < 0.05) reduced by salinity stress, whereas oxidative stress was induced in plants by generating the concentration of reactive oxygen species (ROS) in plant cells/tissues compared to plants grown in the control treatment. Initially, the activity of antioxidant enzymes and relative gene expression increased to a saline level of 100 mM, and then decreased significantly (P < 0.05) by increasing the saline level (150 mM) in the soil compared to plants grown at 0 mM of salinity. We also elucidated that negative impact of salt stress in H. vulgare plants can overcome by the exogenous application of AsA, which not only increased morpho-physiological traits but decreased oxidative stress in the plants by increasing activities of enzymatic antioxidants. We have also explained the negative effect of salt stress on H. vulgare can decrease by exogenous application of AsA, which not only improved morpho-physiological characteristics, ions accumulation in the roots and shoots of the plants, but decreased oxidative stress in plants by increasing antioxidant compounds (enzymatic and non-enzymatic). Taken together, recognizing AsA's role in nutrient uptake introduces new possibilities for agricultural use of this compound and provides a valuable basis for improving plant tolerance and adaptability to potential salinity stress adjustment.     

Metal/metalloid stress tolerance in plants: role of ascorbate,its redox couple,and associated enzymes

The enhanced generation of reactive oxygen species (ROS) under metal/metalloid stress is most common in plants, and the elevated ROS must be successfully metabolized in order to maintain plant growth, development, and productivity. Ascorbate (AsA) is a highly abundant metabolite and a water-soluble antioxidant, which besides positively influencing various aspects in plants acts also as an enigmatic component of plant defense armory. As a significant component of the ascorbate-glutathione (AsA-GSH) pathway, it performs multiple vital functions in plants including growth and development by either directly or indirectly metabolizing ROS and its products. Enzymes such as monodehydroascorbate reductase (MDHAR, EC 1.6.5.4) and dehydroascorbate reductase (DHAR, EC 1.8.5.1) maintain the reduced form of AsA pool besides metabolically controlling the ratio of AsA with its oxidized form (dehydroascorbate, DHA). Ascorbate peroxidase (APX, EC 1.11.1.11) utilizes the reduced AsA pool as the specific electron donor during ROS metabolism. Thus, AsA, its redox couple (AsA/DHA), and related enzymes (MDHAR, DHAR, and APX) cumulatively form an AsA redox system to efficiently protect plants particularly against potential anomalies caused by ROS and its products. Here we present a critical assessment of the recent research reports available on metal/metalloid-accrued modulation of reduced AsA pool, AsA/DHA redox couple and AsA-related major enzymes, and the cumulative significance of these antioxidant system components in plant metal/metalloid stress tolerance.     

Riboflavin spraying impairs the antioxidant defense system but induces waterlogging tolerance in tobacco

Riboflavin, which causes plants to produce reactive oxygen species (ROS) when exposed to light, is an excellent photosensitizer for biocidal reactions. This study explores the possible protective role of riboflavin against waterlogging stress in tobacco plants. Tobacco seedlings (4 weeks old) were divided into four groups and pretreated with 0, 0.2, 0.5 or 1.0 mM riboflavin for 1 week, after which all groups were exposed to waterlogging stress for 7 days. We observed delayed leaf senescence and extended survival time, suggesting that riboflavin can confer increased waterlogging tolerance to plants as compared with the control (0 mM riboflavin). Enhanced stomatal closure was observed in the riboflavin-pretreated tobacco. We evaluated the levels of oxidative damage (H₂O₂ and lipid peroxidation), antioxidant enzyme (superoxide dismutase, catalase, ascorbate peroxidase and glutathione reductase) activity and antioxidant metabolites (including ascorbate and glutathione) in tobacco leaves that were pretreated with riboflavin. However, the results show that riboflavin pretreatment caused a decrease in chlorophyll content, antioxidant enzyme activity and redox values (AsA/DHA and GSH/GSSG), while causing a significant increase in lipid peroxidation, H₂O₂ accumulation and total ascorbate or glutathione content. In addition, the survival time and stomatal aperture of riboflavin-treated plants were significantly modified by exogenous application of GSH, well-known ROS scavenger. To explain the stomatal closure observed in tobacco plants, we propose a “damage avoidance” hypothesis based on riboflavin-mediated ROS toxicity. The protective function of the photosensitizer riboflavin may be highly significant for farming in frequently waterlogged areas.     

Enhancement of antioxidant enzyme activities in rice callus by ascorbic acid under salinity stress

Ascorbic acid (AsA) is naturally occurring compound with antioxidant activity and plays a pivotal role in plant cell adaptation to salinity stress. The objective of this work was to assess the influence of exogenous AsA on the embryogenic callus of indica rice (Oryza sativa L.) cv. MRQ74 cultivated under saline conditions. NaCl (200 mM) decreased callus fresh and dry masses, relative growth rate, and K⁺ and Ca⁺² content, and increased Na⁺ content and Na⁺/K⁺ ratio. Application of AsA (0.5 or 1 mM) alleviated these effects of salinity. Activities of peroxidase, catalase, superoxide dismutase, as well as content of proline increased due to the NaCl treatment, and these parameters were mostly further increased by 0.5 mM AsA. Thus, AsA can increase callus tolerance to NaCl stress.     

Spermidine application enhances tomato seedling tolerance to salinity-alkalinity stress by modifying chloroplast antioxidant systems

The purpose of this study was to elucidate whether exogenous spermidine (Spd) protection of tomato (Solanum lycopersicum L.) seedlings under salinity-alkalinity stress is associated with antioxidant enzymes in the chloroplast. The effects of exogenous Spd on antioxidant enzyme activity and antioxidant content in the chloroplast were evaluated in seedlings of salt-sensitive ecotype (Zhongza 9) grown in a 75 mM salinity-alkalinity solution, with or without 0.25 mM Spd foliar spraying. Results showed that salinity-alkalinity stress increased MDA content, superoxide anion O₂^?- generation rate, superoxide dismutase (SOD), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR) activities and ratio of AsA/DHA and reduced contents of ascorbate (AsA), dehydroascorbate (DHA), AsA+DHA, glutathione (GSH), oxidized glutathione (GSSG), GSH+GSSG, dehydroascorbate reductase (DHAR) activity and ratio of GSH/GSSG in chloroplasts. The exogenous Spd application combined with salinity-alkalinity stress decreased the O₂^?- generation rate and MDA content compared to salinity-alkalinity stress alone. The exogenous Spd also increased AsA-GSH cycle components and increased all antioxidant enzyme activities in most cases. Therefore, exogenous Spd alleviates salinity-alkalinity stress damage using antioxidant enzymes and non-enzymatic systems in chloroplasts.     

外源抗坏血酸对镉胁迫下黑藻抗氧化系统的保护作用

研究了不同浓度抗坏血酸(ascorbic acid,AsA)对5 mg·L^-1Cd²⁺胁迫下黑藻（Hydrilla verticillata）体内超氧阴离子(O₂^-)产生速率、过氧化氢(H₂O₂)含量、抗坏血酸和谷胱甘肽(GSH)含量,以及超氧化物歧化酶(SOD)、超氧化物酶(POD)、过氧化氢酶(CAT)和抗坏血酸过氧化物酶(APX)活性的影响.结果表明,与单一Cd²⁺胁迫相比,随着外源AsA浓度的升高,黑藻体内活性氧生成速率逐步降低;抗氧化物质AsA先升后降,GSH缓慢升高;抗氧化酶APX和CAT活性先升后降,POD活性逐步下降,并接近正常状态,对SOD活性影响不大.可见AsA能够有效缓解Cd²⁺对黑藻的毒害,且60 mg·L^-1浓度下的缓解效果最好.     

Physiological and Biochemical Mechanisms of Exogenous Calcium Chloride on Alleviating Salt Stress in Two Tartary Buckwheat (<i>Fagopyrum tataricum</i>) Varieties Differing in Salinity Tolerance

Salt stress is one of the most serious abiotic stresses limiting plant growth and development. Calcium as an essential nutrient element and important signaling molecule plays an important role in ameliorating the adverse effect of salinity on plants. This study aimed to investigate the impact of exogenous calcium on improving salt tolerance in Tartary buckwheat cultivars, cv. Xinong9920 (salt-tolerant) and cv. Xinong9909 (salt-sensitive). Four-week-old Tartary buckwheat seedlings under 100 mM NaCl stress were treated with and without exogenous calcium chloride (CaCl₂), Ca²⁺ chelator ethylene glycol tetraacetic acid (EGTA) and Ca²⁺-channel blocker lanthanum chloride (LaCl₃) for 10 days. Then, some important physiological and biochemical indexes were determined. The results showed that salt stress significantly reduced seedling growth, decreased photosynthetic pigments, inhibited antioxidants and antioxidant enzyme activities. However, it increased the reactive oxygen species (ROS) levels in the two Tartary buckwheat cultivars. Exogenous 10 mM CaCl₂ application on salt-stressed Tartary buckwheat seedlings obviously mitigated the negative effects of NaCl stress and partially restored seedlings growth. Ca²⁺-treated salt-stressed seedlings diplayed a suppressed accumulation of ROS, increased the contents of total chlorophyll, soluble protein, proline and antioxidants, and elevated the activities of antioxidant enzymes compared with salt stress alone. On the contrary, the addition of 0.5 mM LaCl₃ and 5 mM EGTA on salt-stressed Tartary buckwheat seedlings exhibited the opposite effects to those with CaCl₂ treatment. These results indicate that exogenous Ca²⁺ can enhance salt stress tolerance and Ca²⁺ supplementation may be an effective practice to cultivate Tartary buckwheat in saline soils.     

水孔蛋白在细胞延长、盐胁迫和光合作用中的作用

水孔蛋白属于一个高度保守的、能够进行跨生物膜水分运输的通道蛋白MIP家族。水孔蛋白作为膜水通道,在控制细胞和组织的水含量中扮演重要角色。本研究的重点是属于PIP亚家族的GhPIP1;2和属于TIP亚家族的γTIP1在植物细胞延长中的作用。使用特异基因探针的Northern杂交和实时荧光PCR技术证明GhPIP1;2和GhγTIP1主要在棉花纤维延长过程中显著表达,且最高表达量在开花后5d。在细胞延长过程中,GhPIP1;2和GhγTIP1表达显著,表明它们在促使水流迅速进入液泡这一过程中扮演重要角色。而且也研究了盐胁迫植物中钙离子对水孔蛋白的影响。分别或一起用NaCl或CaCl2处理原生质体或细胞质膜。结果发现在盐胁迫条件下,水渗透率值在原生质体和质膜颗粒中都下降了,同时PIP1水孔蛋白的含量也下降了,表明NaCl对水孔蛋白的功能和含量有抑制作用。同时也观察了Ca2+的两种不同的作用。感知胁迫的胞质中游离钙离子浓度的增加可能导致水孔蛋白的关闭。而过剩的钙离子将导致水孔蛋白的上游调控。同时实验已经证明大麦的一类水孔蛋白-HvPIP2;1有更高的水和CO2转移率。本研究的目标是确定负责转运水和CO2的关键水孔蛋白...     

Manganese-induced salt stress tolerance in rice seedlings: regulation of ion homeostasis,antioxidant defense and glyoxalase systems

Hydroponically grown 12-day-old rice (Oryza sativa L. cv. BRRI dhan47) seedlings were exposed to 150 mM NaCl alone and combined with 0.5 mM MnSO₄. Salt stress resulted in disruption of ion homeostasis by Na⁺ influx and K⁺ efflux. Higher accumulation of Na⁺ and water imbalance under salinity caused osmotic stress, chlorosis, and growth inhibition. Salt-induced ionic toxicity and osmotic stress consequently resulted in oxidative stress by disrupting the antioxidant defense and glyoxalase systems through overproduction of reactive oxygen species (ROS) and methylglyoxal (MG), respectively. The salt-induced damage increased with the increasing duration of stress. However, exogenous application of manganese (Mn) helped the plants to partially recover from the inhibited growth and chlorosis by improving ionic and osmotic homeostasis through decreasing Na⁺ influx and increasing water status, respectively. Exogenous application of Mn increased ROS detoxification by increasing the content of the phenolic compounds, flavonoids, and ascorbate (AsA), and increasing the activities of monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), superoxide dismutase (SOD), and catalase (CAT) in the salt-treated seedlings. Supplemental Mn also reinforced MG detoxification by increasing the activities of glyoxalase I (Gly I) and glyoxalase II (Gly II) in the salt-affected seedlings. Thus, exogenous application of Mn conferred salt-stress tolerance through the coordinated action of ion homeostasis and the antioxidant defense and glyoxalase systems in the salt-affected seedlings.     

外源γ-氨基丁酸对低氧胁迫下甜瓜幼苗活性氧代谢的影响

以甜瓜品种‘西域一号’幼苗为材料,采用营养液水培方法,设置正常通气(对照)、正常通气+GABA(5mmol.L-1)、低氧胁迫、低氧胁迫+GABA(5mmol.L-1)4个处理,研究了外源γ-氨基丁酸(GABA)对正常通气和低氧胁迫下甜瓜幼苗活性氧代谢的影响。结果表明:与正常通气处理相比,低氧胁迫处理导致甜瓜幼苗体内O2.-产生速率和H2O2、MDA含量显著增加,同时SOD、POD、CAT、APX、GR等抗氧化酶活性和抗氧化物质AsA、GSH含量显著提高。低氧胁迫下外源GABA能显著提高甜瓜幼苗叶片SOD、CAT、APX、GR等酶活性和AsA、GSH含量,降低了植株体内O2.-产生速率和H2O2、MDA含量;而正常通气条件下添加外源GABA处理对甜瓜幼苗活性氧代谢的影响较小,仅CAT、GR活性和AsA、GSH含量显著提高,而H2O2、MDA含量显著降低。结果证明,添加外源GABA可以通过显著提高低氧胁迫下抗氧化酶活性和抗氧化物质含量来降低甜瓜幼苗活性氧积累,维持其细胞膜结构稳定性,从而有效减轻低氧胁迫对甜瓜幼苗的伤害。     

Effects of exogenous spermidine on antioxidant system of tomato seedlings exposed to high temperature stress

The effects of foliar spraying with spermidine (Spd) on antioxidant system in tomato (Lycopersicon esculentum Mill.) seedlings were investigated under high temperature stress. The high temperature stress significantly inhibited plant growth and reduced chlorophyll (Chl) content. Application of exogenous 1 mM Spd alleviated the inhibition of growth induced by the high temperature stress. Malondialdehyde (MDA), hydrogen peroxide (H₂O₂) content and superoxide anion (O₂) generation rate were significantly increased by the high temperature stress, but Spd significantly reduced the accumulation of reactive oxygen species (ROS) and MDA content under the stress. The high temperature stress significantly decreased glutathione (GSH) content and activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX), glutathione reductase (GR), monodehydroascorbate reductase (MDHAR) and dehydroascorbate reductase (DHAR), but increased contents of dehydroascorbic acid (DHA), ascorbic acid (AsA), and oxidized glutathione (GSSG) in tomato leaves. However, Spd significantly increased the activities of antioxidant enzymes, levels of antioxidants and endogenous polyamines in tomato leaves under the high temperature stress. In addition, to varying degrees, Spd regulated expression of MnSOD, POD, APX2, APX6, GR, MDHAR, DHAR1, and DHAR2 genes in tomato leaves exposed to the high temperature stress. These results suggest that Spd could change endogenous polyamine levels and alleviate the damage by oxidative stress enhancing the non-enzymatic and enzymatic antioxidant system and the related gene expression.     

Brassinosteroids alleviate chilling-induced oxidative damage by enhancing antioxidant defense system in suspension cultured cells of <Emphasis Type="Italic">Chorispora bungeana</Emphasis>

Brassinosteroids (BRs), a class of plant steroid hormones, play a significant role in the amelioration of various biotic and abiotic stresses. In order to further explore and elaborate their roles in plants subjected to chilling stress, suspension cultured cells of Chorispora bungeana with or without 24-epibrassinolide (EBR) application were exposed to 4 and 0°C for 5 days. The EBR treated cells exhibited higher viability after exposure to low temperatures compared with the control. Under chilling stress, reactive oxygen species (ROS) levels and lipid peroxidation were increased in the cultured cells, which were significantly inhibited by EBR application. The activities of antioxidative enzymes such as ascorbate peroxidase (APX), catalase (CAT), peroxidase (POD) and superoxide dismutase (SOD) were increased during chilling treatments, and these increases were more significant in the EBR applied suspension cells. The EBR treatment also greatly enhanced contents of ascorbic acid (AsA) and reduced glutathione (GSH) under chilling stress. From these results, it can be concluded that EBR could play the positive roles in the alleviation of oxidative damage caused by ROS overproduction through enhancing antioxidant defense system, resulting in improving the tolerance of C. bungeana suspension cultures to chilling stress.     

外源γ-氨基丁酸调控甜瓜叶绿体活性氧代谢应对短期盐碱胁迫

以甜瓜品种‘金辉1号’为试材,采用深液流水培法,研究外源γ 氨基丁酸(GABA)对短期盐碱胁迫下甜瓜幼苗叶绿体活性氧代谢的调控作用.结果表明: 盐碱胁迫显著提高了甜瓜叶绿体内光合色素、丙二醛(MDA)和过氧化氢(H₂O₂)含量及超氧阴离子(O^-_·2)产生速率;增加抗坏血酸(AsA)和谷胱甘肽(GSH)等抗氧化物质含量;明显抑制H⁺-ATP酶(H⁺-ATPase)和H⁺ 焦磷酸酶(H⁺-PPiase)活性.外源叶面喷施GABA有效抑制了盐碱胁迫引起的叶绿体内O^-_·2、H₂O₂和MDA的积累,缓解了光合色素增加的趋势;显著提高SOD和AsA GSH循环各个酶的活性,增加了AsA和GSH库,降低了AsA/DHA和GSH/GSSH比值,增强了H⁺-ATPase和H⁺-PPiase 活性.表明外源GABA能加快叶绿体内活性氧代谢,促进AsA-GSH循环的运转,维持细胞膜的渗透性,进而缓解了盐碱胁迫引起的氧化伤害.     

Salt tolerance of a cash crop halophyte Suaeda fruticosa: biochemical responses to salt and exogenous chemical treatments

Suaeda fruticosa Forssk is a leaf succulent obligate halophyte that produces numerous seeds under saline conditions. Seeds are a good source of high quality edible oil and leaves are capable of removing substantial amount of salt from the saline soil besides many other economic usages. Little is known about the biochemical basis of salt tolerance in this species. We studied some biochemical responses of S. fruticosa to different exogenous treatments under non-saline (0 mM), moderate (300 mM) or high (600 mM) NaCl levels. Eight-week-old seedlings were sprayed twice a week with distilled water, hydrogen peroxide (H₂O₂, 100 μM), glycine betaine (GB, 10 mM), or ascorbic acid (AsA, 20 mM) for 30 days. At moderate (300 mM) NaCl, leaf Na⁺, Ca²⁺ and osmolality increased, along with unchanged ROS and antioxidant enzyme activities, possibly causing a better plant growth. Plants grew slowly at 600 mM NaCl to avoid leaf Na⁺ buildup relative to those at 300 mM NaCl. Exogenous application of distilled water and H₂O₂ improved ROS scavenging mechanisms, although growth was unaffected. ASA and GB alleviated salt-induced growth inhibition at 600 mM NaCl through enhancing the antioxidant defense system and osmotic and ion homeostasis, respectively.     

Plant growth and responses of antioxidants of <Emphasis Type="Italic">Chenopodium album</Emphasis> to long-term NaCl and KCl stress

The effects of long-term NaCl and KCl treatment on plant growth and antioxidative responses were investigated in Chenopodium album, a salt-resistant species widely distributed in semi-arid and light-saline areas of Xinjiang, China. Growth parameters [plant height, branch number, leaf morphology and chlorophyll (Chl) content], the level of oxidative stress [superoxide anion radical (O₂ ⁻), hydrogen peroxide (H₂O₂) and malondialdehyde (MDA) concentrations], activity of antioxidant enzymes [superoxide dismutase (SOD), catalase (CAT), peroxidase (POX)], the contents of non-enzymatic antioxidants [carotenoids (Car) and ascorbic acid (AsA)] and expression of selected genes were investigated. Plants were grown in the presence of 0, 50, and 300 mM NaCl or KCl for 2 months. Growth was stimulated by 50 mM NaCl or KCl, maintained stable at 300 mM NaCl, but was inhibited by 300 mM KCl. Three hundred mM NaCl did not affect O₂ ⁻, H₂O₂, MDA, Car and AsA, but increased the activities of SOD, CAT and POX compared to the controls. RT-PCR analysis suggested that expression of some genes encoding antioxidant enzymes could be induced during long-term salt stress, which was consistent with the enzyme activities. Treatment with 300 mM KCl was associated with elevated oxidative stress, and significantly decreased Car and AsA contents. These results suggest that an efficient antioxidant machinery is important for overcoming oxidative stress induced by treatment with high NaCl concentrations in C. album. Other strategies of ion regulation may also contribute to the differential tolerance to Na and K at higher concentrations.     

Overexpression of dehydroascorbate reductase, but not monodehydroascorbate reductase, confers tolerance to aluminum stress in transgenic tobacco

Aluminum (Al) inhibits plant growth partly by causing oxidative damage that is promoted by reactive oxygen species and can be prevented by improving antioxidant capacity. Ascorbic acid (AsA), the most abundant antioxidant in plants, is regenerated by the action of monodehydroascorbate reductase (MDAR) and dehydroascorbate reductase (DHAR). We investigated the role of MDAR and DHAR in AsA regeneration during Al stress using transgenic tobacco (Nicotiana tabacum) plants overexpressing Arabidopsis cytosolic MDAR (MDAR-OX) or DHAR (DHAR-OX). DHAR-OX plants showed better root growth than wild-type (SR-1) plants after exposure to Al for 2 weeks, but MDAR-OX plants did not. There was no difference in Al distribution and accumulation in the root tips among SR-1, DHAR-OX, and MDAR-OX plants after Al treatment for 24 h. However, DHAR-OX plants showed lower hydrogen peroxide content, less lipid peroxidation and lower level of oxidative DNA damage than SR-1 plants, whereas MDAR-OX plants showed the same extent of damage as SR-1 plants. Compared with SR-1 plants, DHAR-OX plants consistently maintained a higher AsA level both with and without Al exposure, while MDAR-OX plants maintained a higher AsA level only without Al exposure. Also, DHAR-OX plants maintained higher APX activity under Al stress. The higher AsA level and APX activity in DHAR-OX plants contributed to their higher antioxidant capacity and higher tolerance to Al stress. These findings show that the overexpression of DHAR, but not of MDAR, confers Al tolerance, and that maintenance of a high AsA level is important to Al tolerance.     

外源24-表油菜素内酯对高温胁迫下茄子幼苗生长和抗氧化系统的影响

本文研究了高温胁迫下外源24-表油菜素内酯（EBR）对茄子幼苗生长和抗氧化系统的影响。结果表明,外源EBR处理显著促进了高温胁迫下茄子幼苗生长,提高了SOD、POD、CAT和APXS活性,AsA和GSH含量及可溶性蛋白和脯氨酸含量,降低了MDA、O2^-及H2O含量。表明,外源EBR处理通过促进高温胁迫下茄子幼苗抗氧化酶活性、抗氧化剂含量及渗透调节物质的提高,降低ROS水平,缓解高温胁迫对茄子幼苗生长的抑制作用,增强植株抗高温胁迫的能力。     

Methylation of aquaporins in plant plasma membrane

A thorough analysis, using MS, of aquaporins expressed in plant root PM (plasma membrane) was performed, with the objective of revealing novel post-translational regulations. Here we show that the N-terminal tail of PIP (PM intrinsic protein) aquaporins can exhibit multiple modifications and is differentially processed between members of the PIP1 and PIP2 subclasses. Thus the initiating methionine was acetylated or cleaved in native PIP1 and PIP2 isoforms respectively. In addition, several residues were detected to be methylated in PIP2 aquaporins. Lys3 and Glu6 of PIP2;1, one of the most abundant aquaporins in the PM, occurred as di- and mono-methylated residues respectively. Ectopic expression in Arabidopsis suspension cells of PIP2;1, either wild-type or with altered methylation sites, revealed an interplay between methylation at the two sites. Measurements of water transport in PM vesicles purified from these cells suggested that PIP2;1 methylation does not interfere with the aquaporin intrinsic water permeability. In conclusion, the present study identifies methylation as a novel post-translational modification of aquaporins, and even plant membrane proteins, and may represent a critical advance towards the identification of new regulatory mechanisms of membrane transport.