The Photoprotective Role of Spermidine in Tomato Seedlings under Salinity-Alkalinity Stress

Polyamines are small, ubiquitous, nitrogenous compounds that scavenge reactive oxygen species and stabilize the structure and function of the photosynthetic apparatus in response to abiotic stresses. Molecular details underlying polyamine-mediated photoprotective mechanisms are not completely resolved. This study investigated the role of spermidine (Spd) in the structure and function of the photosynthetic apparatus. Tomato seedlings were subjected to salinity-alkalinity stress with and without foliar application of Spd, and photosynthetic and morphological parameters were analyzed. Leaf dry weight and net photosynthetic rate were reduced by salinity-alkalinity stress. Salinity-alkalinity stress reduced photochemical quenching parameters, including maximum photochemistry efficiency of photosystem II, quantum yield of linear electron flux, and coefficient of photochemical quenching (qP). Salinity-alkalinity stress elevated nonphotochemical quenching parameters, including the de-epoxidation state of the xanthophyll cycle and nonphotochemical quenching (NPQ). Microscopic analysis revealed that salinity-alkalinity stress disrupted the internal lamellar system of granal and stromal thylakoids. Exogenous Spd alleviated the stress-induced reduction of leaf dry weight, net photosynthetic rate, and qP parameters. The NPQ parameters increased by salinity-alkalinity stress were also alleviated by Spd. Seedlings treated with exogenous Spd had higher zeaxanthin (Z) contents than those without Spd under salinity-alkalinity stress. The chloroplast ultrastructure had a more ordered arrangement in seedlings treated with exogenous Spd than in those without Spd under salinity-alkalinity stress. These results indicate that exogenous Spd can alleviate the growth inhibition and thylakoid membrane photodamage caused by salinity-alkalinity stress. The Spd-induced accumulation of Z also may have an important role in stabilizing the photosynthetic apparatus.     

Comparative proteomic analysis reveals the positive effect of exogenous spermidine on photosynthesis and salinity tolerance in cucumber seedlings

Key message

Our results based on proteomics data and physiological alterations proposed the putative mechanism of exogenous Spd enhanced salinity tolerance in cucumber seedlings.

Abstract

Current studies showed that exogenous spermidine (Spd) could alleviate harmful effects of salinity. It is important to increase our understanding of the beneficial physiological responses of exogenous Spd treatment, and to determine the molecular responses underlying these responses. Here, we combined a physiological analysis with iTRAQ-based comparative proteomics of cucumber (Cucumis sativus L.) leaves, treated with 0.1 mM exogenous Spd, 75 mM NaCl and/or exogenous Spd. A total of 221 differentially expressed proteins were found and involved in 30 metabolic pathways, such as photosynthesis, carbohydrate metabolism, amino acid metabolism, stress response, signal transduction and antioxidant. Based on functional classification of the differentially expressed proteins and the physiological responses, we found cucumber seedlings treated with Spd under salt stress had higher photosynthesis efficiency, upregulated tetrapyrrole synthesis, stronger ROS scavenging ability and more protein biosynthesis activity than NaCl treatment, suggesting that these pathways may promote salt tolerance under high salinity. This study provided insights into how exogenous Spd protects photosynthesis and enhances salt tolerance in cucumber seedlings.

     

外源亚精胺对根际低氧胁迫下黄瓜幼苗光合作用的影响

采用营养液水培法,研究了低氧胁迫下亚精胺(Spd)对黄瓜幼苗净光合速率（P_n）、细胞间隙CO₂浓度（C_i）、气孔导度（G_s）、蒸腾速率（T_r）以及量子效率（Φ_c）和羧化效率（CE）的影响.结果表明,低氧胁迫下,黄瓜植株P_n呈下降趋势,处理10 d后达最低值,为同期对照的63.33%,而低氧胁迫的外源Spd处理10 d时P_n升高了1.25倍;C_i与P_n呈一定负相关性（R²=0.4730～0.7118）,G_s与T_r的变化幅度较大,低氧胁迫下有明显下降趋势,Spd处理后其值有所上升,两者呈显著相关（R²=0.7821～0.9458）,但与P_n的相关性不显著;低氧下Φ_c和CE比对照分别下降了63.01%和72.33%,而Spd处理后,Φ_c和CE值分别提高了23%和14%.表明在低氧胁迫下黄瓜幼苗的光合抑制主要是由非气孔限制所引起的,而外源Spd可通过对光系统的修饰减轻黄瓜幼苗的低氧胁迫伤害.     

外源亚精胺对NaCl胁迫下毕氏海蓬子光合参数和叶绿体超微结构的影响

在0、100、300、500和700 mmol·L-1NaCl胁迫条件下比较了喷施0.1mmol·L-1亚精胺(Spd)对毕氏海蓬子(Salicomia bigelovii Torr.)幼苗叶绿素含量、净光合速率、气孔导度、胞间CO2浓度和叶绿体超微结构的影响.结果表明:毕氏海蓬子的叶绿素含量、净光合速率和气孔导度均呈低浓度条件下(0、100和300 mmol·L-1NaCl)升高、高浓度条件下(500和700 mmol·L-1NaCl)降低的趋势,在300 mmol·L-1 NaCl胁迫条件下达到最高值:胞间CO2浓度则呈低浓度NaCl胁迫条件下降低、500 mmol·L-1NaCl条件下升高、700 mmol·L-1NaCl条件下略降低的趋势;在0～500 mmol·L-1NaCl胁迫条件下叶绿素a/b值变化不明显,但在700 mmol·L-1NaCl条件下急剧降低.在低浓度NaCl胁迫条件下,叶绿体整体膨胀,类囊体片层结构松散,但叶绿体和类囊体结构仍保持完整;而经500和700mmol·L-1NaCl处理后,叶绿体超微结构被严重破坏,叶绿体膜结构破裂、类囊体结构松散呈放射状、有些叶绿体完全解体.而在相应的NaCl胁迫条件下喷施0.1 mmol·L-1Spd,毕氏海蓬子的叶绿素含量、净光合速率、气孔导度和胞间CO2浓度虽然也呈现出相同的变化趋势,但其数值均显著高于对照(未喷施Spd);且叶绿体超微结构的损伤程度也轻于对照.研究结果说明:喷施外源Spd能够减缓NaCl胁迫对毕氏海蓬子的伤害作用.     

外源多胺对低氧胁迫下黄瓜幼苗根系生长及H+-ATP酶和H+-焦磷酸酶活性的影响

采用营养液水培方式,研究了根际低氧胁迫下外源多胺对黄瓜幼苗植株根系生长,内源多胺含量与质膜H -ATP酶、液泡膜H -ATP酶和焦磷酸酶活性的影响.结果表明,根际低氧胁迫显著抑制黄瓜幼苗根系的生长,外源Put(腐胺)和Spd(亚精胺)可缓解低氧胁迫对根系的生长抑制,多胺主要以Spd的形式发挥促进性的生理作用,Put通过转化为Spd发挥作用;低氧胁迫下黄瓜根系内源多胺含量略有提高,外源多胺处理可增加内源多胺的含量;低氧胁迫下外源Put和Spd处理后质膜H -ATP酶活性显著提高,外源多胺对黄瓜根系液胞膜H -ATP酶和H -焦磷酸酶活性没有明显影响,说明低氧胁迫下外源多胺主要通过提高质膜H -ATP酶活性而发挥生理作用.     

低氧胁迫下24-表油菜素内酯对黄瓜幼苗叶片光合特性及多胺含量的影响

采用1/2 Hoagland营养液培养,研究了低氧胁迫下24-表油菜素内酯(EBR)对黄瓜幼苗叶片光合特性及多胺含量的影响.结果表明：低氧胁迫下黄瓜幼苗的净光合速率(P_n)、气孔导度(g_s)、蒸腾速率(T_r)、胞间CO₂浓度(C_i)显著下降,而叶绿素含量显著提高,幼苗生长受抑;低氧胁迫显著提高了黄瓜幼苗叶片的腐胺(Put)、亚精胺(Spd)、精胺(Spm)、多胺(PAs)含量和Put/PAs,但降低了(Spd+Spm) /Put.低氧胁迫下,外源EBR不仅显著提高了黄瓜幼苗的P_n、g_s、T_r及叶绿素含量,也显著提高了黄瓜幼苗叶片的游离态Spm、结合态Spd、Spm及束缚态Put、Spd、Spm含量,促进了PAs的进一步积累,且降低了Put/PAs,提高了(Spd+Spm)/Put．可见,外源EBR调节了黄瓜幼苗内源多胺含量及形态的变化,维持了较高的光合性能,促进了叶面积和干物质量的显著增加,缓解了低氧胁迫对黄瓜幼苗的伤害.     

外源褪黑素对高温胁迫下菊花光合和生理特性的影响

以切花菊品种‘神马'为试材,研究外源褪黑素(MT)对菊花抗高温胁迫的影响。将供试菊花叶面喷施200 μmol·L^-1的MT后,进行40 ℃(昼)/35 ℃(夜)高温胁迫,观察菊花叶片叶绿体和内囊体超微结构,测定光合和生理指标。结果表明:与常温对照(CK)相比,高温胁迫下菊花叶片叶绿体和类囊体受损,叶绿素含量和最大荧光(F_m)显著降低,OJIP曲线发生变化,K点和J点荧光升高,净光合速率(P_n)、蒸腾速率(T_r)、气孔导度(g_s)均显著降低,胞间CO₂浓度(C_i)均显著增加;相对电导率(REC)、丙二醛(MDA)、活性氧(ROS)、渗透调节物质含量和抗氧化酶活性显著增加。外源喷施MT可维持高温胁迫下植株叶绿体和类囊体结构的完整性,明显降低OJIP曲线中K点和J点上升的幅度,F_m、P_n、g_s、T_r和光合色素含量显著提高,C_i显著下降,缓解了高温胁迫对菊花光合和荧光作用的抑制。同时,外源喷施MT处理显著降低了高温胁迫下菊花REC、MDA和ROS含量,增强了菊花叶片中渗透调节物质含量和抗氧化酶活性。可见,外源MT可通过保护菊花叶片叶绿体结构的完整性,增强光合作用,抑制高温胁迫下菊花植株体内ROS的过度产生,提高抗氧化酶系统的活性,降低膜质过氧化水平和保护脂膜的完整性,从而提高菊花植株抗高温胁迫能力。     

外源脯氨酸对高温胁迫下黄瓜幼苗叶片AsA-GSH循环和光合荧光特性的影响

以抗热性较弱的黄瓜品种‘新泰密刺'为试材,在人工气候箱内采用营养液栽培法,研究了外源脯氨酸(Pro)预处理对高温胁迫下黄瓜幼苗叶片抗坏血酸-谷胱甘肽循环和光合荧光特性的影响.结果显示:(1)与清水处理相比,高温胁迫4 h和8 h时,Pro预处理黄瓜幼苗叶片单脱氢抗坏血酸还原酶(MDAR)活性、GSH(还原型谷胱甘肽)/GSSG(氧化型谷胱甘肽)比值及GSH含量显著升高;(2)在高温胁迫8 h时,Pro预处理幼苗的净光合速率(P_n)、气孔导度(G_s)及PSⅡ的最大光化学效率(F_v/F_m)、光化学淬灭系数(q_P)均显著升高,而蒸腾速率(T_r)和非光化学淬灭系数(NPQ)降低.研究表明,外源Pro预处理可显著提高高温胁迫下黄瓜幼苗叶片抗坏血酸-谷胱甘肽循环清除H_2O_2能力和叶片光合能力,有效缓解高温胁迫对黄瓜叶片抗氧化系统和光合系统的伤害,从而增强植株的耐热性.     

Metabolic regulation of polyamines and γ‐aminobutyric acid in relation to spermidine‐induced heat tolerance in white clover

• Heat stress decreases crop growth and yield worldwide. Spermidine (Spd) is a small aliphatic amine and acts as a ubiquitous regulator for plant growth, development and stress tolerance.
• Objectives of this study were to determine effects of exogenous Spd on changes in endogenous polyamine (PA) and γ‐aminobutyric acid (GABA) metabolism, oxidative damage, senescence and heat shock protein (HSP) expression in white clover subjected to heat stress. Physiological and molecular methods, including colorimetric assay, high performance liquid chromatography and qRT‐PCR, were applied.
• Results showed that exogenous Spd significantly alleviated heat‐induced stress damage. Application of Spd not only increased endogenous putrescine, Spd, spermine and total PA accumulation, but also accelerated PA oxidation and improved glutamic acid decarboxylase activity, leading to GABA accumulation in leaves under heat stress. The Spd-pretreated white clover maintained a significantly higher chlorophyll (Chl) content than untreated plants under heat stress, which could be related to the roles of Spd in up‐regulating genes encoding Chl synthesis (PBGD and Mg‐CHT) and maintaining reduced Chl degradation (PaO and CHLASE) during heat stress. In addition, Spd up‐regulated HSP70, HSP70B and HSP70‐5 expression, which might function in stabilizing denatured proteins and helping proteins to folding correctly in white clover under high temperature stress.
• In summary, exogenous Spd treatment improves the heat tolerance of white clover by altering endogenous PA and GABA content and metabolism, enhancing the antioxidant system and HSP expression and slowing leaf senescence related to an increase in Chl biosynthesis and a decrease in Chl degradation during heat stress.

     

A Comparison of the Effects of Chilling on Thylakoid Electron Transfer in Pea (Pisum sativum L.) and Cucumber (Cucumis sativus L.)

Experiments comparing the photosynthetic responses of a chilling-resistant species (Pisum sativum L. cv Alaska) and a chilling-sensitive species (Cucumis sativus L. cv Ashley) have shown that cucumber photosynthesis is adversely affected by chilling temperatures in the light, while pea photosynthesis is not inhibited by chilling in the light. To further investigate the site of the differential response of these two species to chilling stress, thylakoid membranes were isolated under various conditions and rates of photosynthetic electron transfer were determined. Preliminary experiments revealed that the integrity of cucumber thylakoids from 25°C-grown plants was affected by the isolation temperature; cucumber thylakoids isolated at 5°C in 400 millimolar NaCl were uncoupled, while thylakoids isolated at room temperature in 400 millimolar NaCl were coupled, as determined by addition of gramicidin. The concentration of NaCl in the homogenization buffer was found to be a critical factor in the uncoupling of cucumber thylakoids at 5°C. In contrast, pea thylakoid membranes were not influenced by isolation temperatures or NaCl concentrations. In a second set of experiments, thylakoid membranes were isolated from pea and cucumber plants at successive intervals during a whole-plant light period chilling stress (5°C). During wholeplant chilling, thylakoids isolated from cucumber plants chilled in the light were uncoupled even when the membranes were isolated at warm temperatures. Pea thylakoids were not uncoupled by the whole-plant chilling treatment. The difference in integrity of thylakoid membrane coupling following chilling in the light demonstrates a fundamental difference in photosynthetic function between these two species that may have some bearing on why pea is a chilling-resistant plant and cucumber is a chilling-sensitive plant.     

Glycine betaine improves thylakoid membrane function of tobacco leaves under low-temperature stress

Glycine betaine (GB) is an effective compatible solute that improves the tolerance in plants to various stresses. We investigated the effects of 2 mM GB applied to the roots of a tobacco (Nicotiana tabacum L.) cultivar on enhancing photosynthesis under low-temperature (LT) stress (5/5 °C, 12/12 h, 300 μmol m⁻² s⁻¹) and in the subsequent recovery (25/18 °C) from the stress. The net photosynthetic rate, intrinsic efficiency measured as the ratio of variable to maximum fluorescence, and actual efficiency of the photochemistry of photosystem 2 as well as the ATPase activity in the thylakoid membrane decreased, and a distinct K step in the fluorescence transient O-J-I-P appeared under cold stress. Exogenous GB alleviated the decrease in all these parameters. The LT-stress induced the accumulation of 33–66 kDa polypeptides and decreased the proportion of unsaturated fatty acids in the thylakoid membrane. In plants subjected to LT-stress, GB protected these polypeptides from damage and enhanced the proportion of unsaturated fatty acids. An increase in non-radiative energy dissipation (NPQ) may be involved in the improvement of the function of the thylakoid membrane by GB since exogenous GB protected violaxanthin de-epoxidase and enhanced NPQ.     

外源多胺对低氧胁迫下黄瓜幼苗光合特性和膜脂过氧化的影响

以抗低氧能力不同的2个黄瓜(Cucumis sativus)品种为试材, 研究了外源多胺对黄瓜幼苗植株生长、光合特性和膜脂过氧化的影响。结果表明, 外源多胺能显著提高低氧胁迫下黄瓜幼苗叶片的净光合速率和水分利用率, 降低叶片中丙二醛含量和质膜透性, 使幼苗鲜重和干重明显增加。因此在低氧胁迫下, 外源喷施多胺能提高幼苗叶片的净光合速率, 促进植株生长, 缓解胁迫对黄瓜幼苗的伤害。此外, 与抗低氧能力较强的品种绿霸春4号相比, 外源多胺对抗低氧能力较弱的品种中农8号的影响更明显。     

Effects of salt stress on the structure and function of the photosynthetic apparatus in Cucumis sativus and its protection by exogenous putrescine

With the objective to clarify the physiological significance of polyamines (PAs) in the photosynthetic apparatus, the present study investigated the effects of salt stress with and without foliar application of putrescine (Put) on the structure and function of the photosynthetic apparatus in cucumber. Salt stress at 75 mM NaCl for 7 days resulted in a severe reduction of photosynthesis. The fast chlorophyll afluorescence transient analysis showed that salt stress inhibited the maximum quantum yield of PSII photochemistry (F(v) /F(m) ), mainly due to damage at the receptor side of PSII. In addition, salt stress decreased the density of active reaction centers and the structure performance. The microscopic analysis revealed that salt stress-induced destruction of the chloroplast envelope and increased the number of plastoglobuli along with aberrations in thylakoid membranes. Besides, salt stress caused a decrease in the content of endogenous PAs, conjugated and bound forms of spermidine and spermine in particular, in thylakoid membranes. However, applications of 8 mM Put alleviated the salt stress-mediated decrease in net photosynthetic rates (Pn) and actual efficiency of PSII (Φ(PSII) ). Put increased PAs in thylakoid membranes and overcame the damaging effects of salt stress on the structure and function of the photosynthetic apparatus in salt-stressed plant leaves. Put application to control plants neither increased PAs in thylakoid membranes nor affected photosynthesis. These results indicate that PAs in chloroplasts play crucial roles in protecting the thylakoid membranes against the deleterious influences of salt stress. In addition, the present results point to the probability that the salt-induced dysfunction of photosynthesis is largely attributable to the loss of PAs in the photosynthetic apparatus.     

低温胁迫对类囊体膜脂代谢的影响

类囊体膜主要由膜脂、膜蛋白及一些光合色素等成分组成,它是植物进行光合作用的场所。低温能通过影响类囊体膜的结构而影响植物的光合作用。简述了类囊体膜的组成和功能,以及低温胁迫下类囊体膜脂及其脂肪酸组成的变化。简要介绍了膜脂与光抑制的关系,以及利用分子生物学手段研究三烯脂肪酸与植物抗冷性关系的相关进展。     

外源多胺对低氧胁迫下黄瓜幼苗光合特性和膜脂过氧化的影响

以抗低氧能力不同的2个黄瓜（Cucumis sativus）品种为试材,研究了外源多胺对黄瓜幼苗植株生长、光合特性和膜脂过氧化的影响.结果表明,外源多胺能显著提高低氧胁迫下黄瓜幼苗叶片的净光合速率和水分利用率,降低叶片中丙二醛含量和质膜透性,使幼苗鲜重和干重明显增加.因此在低氧胁迫下,外源喷施多胺能提高幼苗叶片的净光合速率,促进植株生长,缓解胁迫对黄瓜幼苗的伤害.此外,与抗低氧能力较强的品种绿霸春4号相比,外源多胺对抗低氧能力较弱的品种中农8号的影响更明显.     

Environmentally modulated phosphorylation and dynamics of proteins in photosynthetic membranes

Recent advances in vectorial proteomics of protein domains exposed to the surface of photosynthetic thylakoid membranes of plants and the green alga Chlamydomonas reinhardtii allowed mapping of in vivo phosphorylation sites in integral and peripheral membrane proteins. In plants, significant changes of thylakoid protein phosphorylation are observed in response to stress, particularly in photosystem II under high light or high temperature stress. Thylakoid protein phosphorylation in the algae is much more responsive to the ambient redox and light conditions, as well as to CO₂ availability. The light-dependent multiple and differential phosphorylation of CP29 linker protein in the green algae is suggested to control photosynthetic state transitions and uncoupling of light harvesting proteins from photosystem II under high light. The similar role for regulation of the dynamic distribution of light harvesting proteins in plants is proposed for the TSP9 protein, which together with other recently discovered peripheral proteins undergoes specific environment- and redox-dependent phosphorylation at the thylakoid surface. This review focuses on the environmentally modulated reversible phosphorylation of thylakoid proteins related to their membrane dynamics and affinity towards particular photosynthetic protein complexes.