Nitrogen Modulates the Effects of Heat,Drought, and Combined Stresses on Photosynthesis,Antioxidant Capacity,Cell Osmoregulation,and Grain Yield in Winter Wheat

Longer and more intense heat and drought stresses will occur in terrestrial ecosystems in the future. Although the effects of individual heat or drought stress on wheat plants have been largely explored, the regulatory effect of nitrogen (N) on winter wheat under heat, drought, and combined stresses and whether N alleviates damage to wheat plants caused by these stresses remain unclear. Therefore, the objective of the present study was to investigate the growth, photosynthesis, antioxidant enzyme and N metabolism-related enzyme activity, cell membrane system, osmoregulatory substance, and yield responses to heat, drought, and combined stresses in wheat plants and to clarify the regulatory effects of N on the growth, physiological and biochemical characteristics, and yield of wheat plants under stress conditions. The results showed that wheat plant exposure to individual heat or drought stress reduced photosynthesis and N metabolism-related enzyme activities and increased antioxidant enzyme activities, electrolyte leakage (EL), and the contents of MDA (malondialdehyde) and O₂^? (superoxide anion). The above parameters showed typical superposition effects under combined stress. Under individual heat or drought stress, wheat plants treated with a medium (N₂) or high (N₃) N supply maintained higher photosynthesis and N metabolism-related enzyme activities than did those treated with a low N supply (N₁). Enhanced heat and drought tolerance in wheat plants under an appropriate N supply may be attributed to improved antioxidant capacity, as exemplified by increased activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), glutathione reductase (GR) and ascorbate peroxidase (APX), and to enhanced osmoregulation capacity, as signified by increased contents of soluble sugar (SS), soluble protein (SP), and proline (Pro). Variable importance in projection (VIP) analysis indicated that efficient SOD, POD, CAT, and GR activities and an increased Pro content had superior potential to alleviate heat, drought, and combined stress stresses in wheat plants, and the improvements in growth and grain yield in wheat plants further confirmed the oxidative stress alleviation and stress tolerance enhancement. However, positive effects of N on wheat growth and grain yield under combined stress were usually observed under a low N supply. These results may facilitate future research on the effects of N fertilizer on the stress resistance of winter wheat.

Graphical Abstract

     

Comparative effect of water, heat and light stresses on photosynthetic reactions in Sorghum bicolor(L.) Moench

Five varieties of Sorghum bicolor (L.) Moench., differing in their drought tolerance under field conditions have been used to study the effect of individual components of drought stress, namely high light intensity stress, heat stress and water stress, on their photosynthetic performance. Chlorophyll content, chlorophyll fluorescence, ribulose-1,5-bisphosphate carboxylase (Rubisco, EC 4.1.1.39) content, phosphoenolpyruvate carboxylase (PEPcase, EC 4.1.1.31) activity and photo-synthetic oxygen evolution were used as key parameters to assess photosynthetic performance. The results indicated that photochemical efficiency of photosystem II (PSII) was severely reduced by all three stress components, whereas PEPcase activity was more specifically reduced by water stress. Degradation of Rubisco and chlorophyll loss occurred under high light and water stress conditions. Of the four drought-tolerant varieties, E 36-1 showed higher PEPcase activity, Rubisco content and photochemical efficiency of PSII, and was able to sustain a higher maximal rate of photosynthetic oxygen evolution under each stress condition as compared to the other varieties. A high stability to stress-induced damage, or acclimation of photosynthesis to the individual components of drought stress may contribute to the high yields of E 36-1 under drought conditions. In the E 36-1 variety markedly higher levels of the chloroplastic chaperonin 60 (cpn 60) were observed under all stress conditions than in the susceptible variety CSV 5.Key words: Chlorophyll fluorescence, drought stress, oxygen evolution, phosphoenopyruvate carboxylase, Sorghum.      

水分胁迫下不同抗旱性小麦品种叶片转录因子表达差异研究

通过研究不同抗旱性小麦品种中转录因子表达水平的差异,为阐明小麦抗旱机制奠定基础。依据候选基因序列设计PCR引物,以干旱胁迫后0、3、6、9、12和24 h的小麦叶片为实验材料,以26S rRNA为内参,运用荧光定量PCR技术,检测Wdreb2、Wlip19基因在干旱敏感性和干旱耐受性小麦叶片中的相对表达量。定量PCR结果显示：干旱胁迫后,Wdreb2、Wlip19基因在干旱敏感性小麦叶片中的表达明显低于干旱耐受性小麦,在不同品种叶片中的响应时间和表达趋势存在差异。研究认为,Wdreb2、Wlip19基因在不同品种小麦受到干旱胁迫后的表达差异,与该品种小麦的抗旱能力具有一定的相关性。     

Performance of wheat crops with different chromosome ploidy: root-sourced signals, drought tolerance, and yield performance

Pot-culture experiments were carried out to estimate the role of non-hydraulic root signals (nHRS) and the relation of these signals to drought tolerance and grain yield formation under drought stress in six wheat varieties. These were two modern hexaploid wheat (Triticum aestivum L., AABBDD) Plateau602 and Longchun8139-2, two diploid wheat (Triticum monococcum L., AB) MO1 and MO4, and two tetraploid wheat (Triticum dicoccum Schuebl L., AABB) DM22 and DM31. In the two diploid relatives, the nHRS was switched on and off at a soil water content (SWC) of approximately 53–45% field water capacity (FWC). In contrast, in the modern hexaploid varieties, Longchun8139-2 and Plateau602 the nHRS occurred between a SWC of about 71 and 35% FWC, a much wider soil moisture range. The two tetraploid relatives, DM22 and DM31, were generally intermediate. The nHRS threshold range in SWC also narrowed as all six varieties went through successive developmental stages from shooting to grain filling. The two hexaploid wheat varieties had the longest duration of survival after the water supply ceased, and the best yield stability under drought stress, similar to with tetraploid wheat varieties; the diploid wheat varieties were least robust. These two parameters were both significantly correlated with the nHRS soil moisture threshold range (r=0.9456** and 0.8608*, respectively). Based on these patterns, we propose a ‘triple Z’ model to describe the features of non-hydraulic stomatal sensitivity versus soil drought in wheat growth.     

Combined drought and heat stress in wheat: changes in some heat shock proteins

The influence of combined and individually applied drought and heat stress was studied in two wheat (Triticum aestivum L.) cultivars: resistant cv. Katya and susceptible cv. Sadovo. Relative water content decreased and electrolyte leakage increased due to individual and combined application of both stresses. Initial heat shock protein profile has been outlined via SDS electrophoresis of leaf extracts. The results obtained were confirmed by immunoblotting with anti-HSP70 monoclonal antibodies, anti-HSP110 polyclonal antibodies and anti-α β-crystalline polyclonal antibodies. The effect of simultaneously applied water stress and heat shock resembled the alterations in protein expression provoked only by water stress and differed significantly from the changes occurring after the individual application of heat stress.     

Heat stress results in loss of chloroplast Cu/Zn superoxide dismutase and increased damage to Photosystem II in combined drought‐heat stressed Lotus japonicus

Drought and heat stress have been studied extensively in plants, but most reports involve analysis of response to only one of these stresses. Studies in which both stresses were studied in combination have less commonly been reported. We report the combined effect of drought and heat stress on Photosystem II (PSII) of Lotus japonicus cv. Gifu plants. Photochemistry of PSII was not affected by drought or heat stress alone, but the two stresses together decreased PSII activity as determined by fluorescence emission. Heat stress alone resulted in degradation of D1 and CP47 proteins, and D2 protein was also degraded by combined drought–heat stress. None of these proteins were degraded by drought stress alone. Drought alone induced accumulation of hydrogen peroxide but the drought–heat combination led to an increase in superoxide levels and a decrease in hydrogen peroxide levels. Furthermore, combined drought–heat stress was correlated with an increase in oxidative damage as determined by increased levels of thiobarbituric acid reactive substances. Heat also induced degradation of chloroplast Cu/Zn superoxide dismutase (SOD: EC 1.15.1.1) as shown by reduced protein levels and isozyme‐specific SOD activity. Loss of Cu/Zn SOD and induction of catalase (CAT: EC 1.11.1.6) activity would explain the altered balance between hydrogen peroxide and superoxide in response to drought vs combined drought–heat stress. Degradation of PSII could thus be caused by the loss of components of chloroplast antioxidant defence systems and subsequent decreased function of PSII. A possible explanation for energy dissipation by L. japonicus under stress conditions is discussed.     

Changes in the photosynthetic efficiency of winter wheat in response to abiotic stress

The assessment of heat and drought tolerance is of primary importance in breeding programmes designed to improve heat and drought tolerance in cereals. Three winter wheat varieties grown in controlled growth chambers were exposed to heat (H) and drought (D) stress singly and in combination (H+D). The combined effects of H and D stress were much more severe than those of individual treatments for both physiological and yield parameters during grain filling. The chlorophyll content, effective quantum yield of PSII, net assimilation rate, transpiration, stomatal conductance and intercellular CO₂ concentration were greatly reduced by H, D and their interaction. Grain yield decreased to a greater extent (48.3%) in Plainsman V, averaged over the stress treatments, than in Mv Magma (67.8%) and Fatima 2 (53.7%). The least decline was found in grain number, except in Plainsman V. Mv Magma tolerated heat stress better than Fatima 2. In terms of photosynthetic activity, Plainsman V showed better drought tolerance than Mv Magma. The results showed that changes in physiological properties during stress treatment are not always associated with changes in yield parameters, so a combination of methods may be needed to give a more precise picture of the stress tolerance of wheat varieties.     

盐旱复合胁迫对小麦幼苗生长和水分吸收的影响

为明确盐害、干旱及盐旱复合胁迫对小麦幼苗生长和水分吸收的影响,从而为盐害和干旱胁迫下栽培调控提供理论依据。以2个抗旱性不同的小麦品种(扬麦16和耐旱型洛旱7号)为材料,采用水培试验,以NaCl和PEG模拟盐旱复合胁迫,研究了盐旱复合胁迫下小麦幼苗生长、根系形态、光合特性及水分吸收特性的变化。结果表明,盐、旱及复合胁迫下小麦幼苗的生物量、叶面积、总根长与根系表面积、叶绿素荧光和净光合速率均显著下降,但是复合胁迫处理的降幅却显著低于单一胁迫。盐旱复合胁迫下根系水导速率和根系伤流液强度显著大于单一胁迫,从而提高了小麦幼苗叶片水势和相对含水量。盐胁迫下小麦幼苗Na~+/K~+显著大于复合胁迫,但复合胁迫下ABA含量却显著小于单一的盐害和干旱胁迫。因此,盐旱复合胁迫可以通过增强根系水分吸收及降低根叶中ABA含量以维持较高光合能力,这是盐旱复合胁迫提高小麦适应性的重要原因。洛旱7号和扬麦16对盐及盐旱复合胁迫的响应基本一致,但在干旱胁迫下洛旱7号表现出明显的耐性。     

水分胁迫下内生真菌球毛壳ND35对冬小麦苗期生长和抗旱性的影响

为明确不同水分条件下内生真菌对冬小麦苗期生长和抗旱性的影响,以抗旱型小麦品种山农16和水分敏感型小麦品种山农22为材料,利用荧光定量PCR技术检测小麦干旱诱导基因脱水素wzy2的表达量来了解冬小麦在干旱胁迫下相关基因的表达差异,通过测定相关生理指标与酶活性来判断小麦发育及其在干旱胁迫下的生理响应状况。结果表明,与正常水分ND35组相比,接种球毛壳菌(Chaetomium globosum)ND35的干旱处理组小麦的根冠比、总蛋白含量、脯氨酸含量及丙二醛含量等指标显著提高,小麦叶片含水量和可溶性糖含量有所降低。在干旱处理组中,球毛壳菌ND35可以显著提高小麦山农16的根长和山农22的株高,接种球毛壳ND35的山农16脯氨酸含量、可溶性糖含量、过氧化氢酶活性比对照组均显著提高,丙二醛含量比对照组降低9.0%,但差异不显著;山农22脯氨酸含量和过氧化氢酶活性比对照组显著提高,丙二醛含量和可溶性糖含量比对照组有所降低,但可溶性糖含量差异不显著;相对定量检测数据显示,接种球毛壳ND35后,两种小麦脱水素wzy2基因的表达量较对照组均能够显著提高。综合分析说明内生真菌球毛壳ND35可以促进冬小麦苗期根系和植株发育,小麦提前进入三叶期,增强小麦避旱性,同时提高小麦根系活力,增强小麦耐旱性;提高个体细胞内水分、糖分、脯氨酸含量,降低丙二醛的氧化性损伤,增强过氧化氢酶活性,从而提高两种冬小麦对干旱胁迫的耐受能力;球毛壳ND35促进小麦干旱诱导相关基因wzy2的表达量,进而提高抗旱相关蛋白的表达,从而提高两种冬小麦耐脱水性和对干旱胁迫的适应性。     

Overaccumulation of glycine betaine enhances tolerance to drought and heat stress in wheat leaves in the protection of photosynthesis

We investigated the different responses of wheat (Triticum aestivum L.) plants to drought- (DS) and heat stress (HS), and analyzed the physiological mechanisms of glycine betaine (GB) involved in the improvement of wheat tolerance to the combination of these stresses. The transgenic wheat T6 line was generated by introducing a gene encoding betaine aldehyde dehydrogenase (BADH) into the wild-type (WT) Shi4185 line. The gene was cloned from the Garden Orache plant (Atriplex hortensis L.). Wheat seedlings were subjected to drought stress (30%, PEG-6000), heat stress (40°C), and their combination. Photosynthetic gas exchange, water status and lipid peroxidation of wheat leaves were examined under different stresses. When subjected to a combination of drought and heat, the inhibition of photosynthesis was significantly increased compared to that under DS or HS alone. The increased inhibition of photosynthesis by the combined stresses was not simply the additive stress effect of separate heat- and drought treatments; different responses in plant physiology to DS and HS were also found. HS decreased the chlorophyll (Chl) content, net photosynthetic rate (P _N), carboxylation efficiency (CE) and apparent quantum yield (AQY) more than DS but DS decreased the transpiration rate (E), stomata conductance (g _s) and intercellular CO₂ concentration (C _i) more than HS. GB over-accumulation led to increased photosynthesis not only under individual DS or HS but also under their combination. The enhancement of antioxidant activity and the improvement of water status may be the mechanisms underlying the improvement of photosynthesis by GB in wheat plants.     

Improvement of heat and drought photosynthetic tolerance in wheat by overaccumulation of glycinebetaine

Within their natural habitat, crops are often subjected to drought and heat stress, which suppress crop growth and decrease crop production. Causing overaccumulation of glycinebetaine (GB) has been used to enhance the crop yield under stress. Here, we investigated the response of wheat (Triticum aestivum L.) photosynthesis to drought, heat stress and their combination with a transgenic wheat line (T6) overaccumulating GB and its wild-type (WT) Shi4185. Drought stress (DS) was imposed by controlling irrigation until the relative water content (RWC) of the flag leaves decreased to between 78 and 82%. Heat stress (HS) was applied by exposing wheat plants to 40°C for 4 h. A combination of drought and heat stress was applied by subjecting the drought-stressed plants to a heat stress as above. The results indicated that all stresses decreased photosynthesis, but the combination of drought and heat stress exacerbated the negative effects on photosynthesis more than exposure to drought or heat stress alone. Drought stress decreased the transpiration rate (Tr), stomatal conductance (Gs) and intercellular CO₂ concentration (Ci), while heat stress increased all of these; the deprivation of water was greater under drought stress than heat stress, but heat stress decreased the antioxidant enzyme activity to a greater extent. Overaccumulated GB could alleviate the decrease of photosynthesis caused by all stresses tested. These suggest that GB induces an increase of osmotic adjustments for drought tolerance, while its improvement of the antioxidative defense system including antioxidative enzymes and antioxidants may be more important for heat tolerance.     

Rubisco and some chaperone protein responses to water stress and rewatering at early seedling growth of drought sensitive and tolerant wheat varieties

Four wheat varieties differing in their drought tolerance were subjected to severe but recoverable water stress at seedling stage. Growth parameters, leaf water deficit (WD) and electrolyte leakage (EL) were used to evaluate the stress intensity and the extent of recovery. The physiological response of the varieties was quite similar under severe drought. Leaf protein patterns and levels of some individual proteins relevant to ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) maintenance were studied in control, stressed and recovering plants by electrophoresis and immunoblotting. The bands representing Rubisco large subunit (RLS), N- and C-terminus of RLS, Rubisco activase (RA) and Rubisco binding protein (RBP, cpn 60), as well as the chaperone and proteolytic subunits of the Clp protease complex were identified using polyclonal antibodies. Under drought conditions RLS, Clp proteases and especially RBP were enhanced, whereas the RA band was only slightly affected. The drought tolerant varieties had higher RBP content in the controls and drought treated plants. Its concentration could be a potential marker for drought tolerance.     

Genotypic variation in drought stress response and subsequent recovery of wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.)

Three wheat (Triticum aestivum L.) genotypes, Sadovo, Katya and Prelom, with different tolerance to drought were comparatively evaluated in terms of leaf respiratory responses to progressing dehydration and consecutive rewatering. Under drought stress, the respiration of all varieties gradually decreased, as the drought-tolerant Katya showed the most pronounced decline at earlier stages of dehydration. When water stress intensified, this genotype gave relatively stable respiration rates compared with the drought-sensitive varieties. Additionally, dehydrated Katya leaves displayed lower stomatal conductance and higher photosynthesis values, which resulted in greater water use efficiency during the dehydration period. Combination of drought stress and short-term changes in leaf temperature also induced genotype-specific response that differed from the response to drought only. Over the whole temperature range, the leaves of Katya exposed to dehydration for 14 days, showed higher respiration rates compared to the drought-sensitive varieties. The sensitive varieties maintained higher respiration rates under control conditions and mild dehydration, and very low rates under severe drought. In Katya, respiration and photosynthesis were fully restored from the stress within the first day of rewatering. The drought-sensitive genotypes displayed a considerably slower recovering capacity. The results are discussed in terms of possible physiological mechanisms underlying plant tolerance to drought.     

Improved drought resistance in a wheat stay-green mutant tasg1 under field conditions

We investigated the drought resistance of a wheat (Triticum aestivum L.) stay-green mutant tasg1 and its wild-type (WT) in field experiments conducted for two years. Drought stress was imposed by controlling irrigation and sheltering the plants from rain. Compared with the WT, tasg1 exhibited a distinct delayed senescence under both normal and drought stress conditions, as indicated by slower degradation of chlorophyll and decrease in net photosynthetic rate than in WT. At the same time, tasg1 mutants maintained more integrated chloroplasts and thylakoid ultrastructure than did WT plants under drought stress. Lower malondialdehyde content and higher antioxidative enzyme activities in tasg1, compared to WT, may be involved in the stay-green phenotype and drought resistance of tasg1.     

Effect of Pre-Anthesis Drought Hardening on Post-Anthesis Physiological Characteristics,Yield and WUE in Winter Wheat

A drought event can cause yield loss or entire crops to fail. In order to study the effects of continuous drought on physiological characteristics, yield, and water use efficiency (WUE) of winter wheat (Triticum aestivum L.), the variety “Zhoumai 22” was grown in controlled water conditions of the pot-planted winter wheat under a mobile rainout shelter. Foot planting and safe wintering were used to evaluate, winter wheat under different drought conditions, including light, moderate and severe drought at the jointing, heading, and filling stages. The soil water content was controlled at 60–70%, 50–60%, or 40–50% of field capacity. Experimental trials included 3 pre-anthesis drought hardening, 3 three-stage continuous drought, and 1 test control conditions. Under drought stress conditions, winter wheat leaf water potential, soil plant analysis development (SPAD), photosynthesis parameters, and yield declined due to pre-anthesis drought hardening. And the degree of decline: continuous drought > pre-anthesis drought hardening. Changes in the post-anthesis photosynthetic capacity of winter wheat were mainly related to the pre-anthesis drought level, rather than post-anthesis rehydration. The threshold of non-stomata limiting factors caused by photosynthesis at the filling stage is 40–50%FC, while comprehensive yield and WUE affected, the yield in severe drought treatments was the most significant, B3C3 and B3C3G3 decreased by 55.68% and 55.88%, respectively. Pre-anthesis drought was the main reason for the decreased crop yield. Thus, severe drought should be avoided during planting, while pre-anthesis light drought is a suitable choice for water-saving and crop production, as proper pre-anthesis drought hardening (60–70% FC) is feasible and effective.     

Physiological response of tomatoes at drought,heat and their combination followed by recovery

In nature, crops encounter a combination of abiotic stresses that severely limit yield. Our aim was to dynamically expose the changes of tomatoes' physiological parameters to drought, heat and their combination and thereby clarify the relationship between the responses to single and combined stress. We studied the effect of single and combined drought and heat stresses on the shoot and root of two tomato cultivars (Sufen No.14 as CV1; Jinlingmeiyu as CV2). After being exposed to combined stress for 6 days, the dry weight of shoot and root significantly decreased. The F_q′/F_m′ (quantum yield of photosystem II) was significantly lower in CV1 upon drought and combined stress and in CV2 subjected to combined stress (between days 4 and 6) compared to control. The relative water content during combined stress was significantly lower than control from day 4 to recovery day 2. On days 3 and 6, the water loss rate significantly increased under heat stress and decreased at drought and combined stress, respectively. The combined stress caused severe damages on photosystem II and chloroplast ultrastructure. The root activity after stress recovered even though drought significantly increased the activity from day 2 to day 6. Combined stress result in complex responses during tomato growth. The CV1 was more heat tolerant than CV2, but there was no varietal difference at drought and combined stress. This study contributes to the understanding of the underlying physiological response mechanism of plant to combined stress and crop improvement by providing valuable information for abiotic stress‐tolerant tomato breeding.     

<Emphasis Type="Bold">Role of microRNAs and their target genes in salinity response in plant</Emphasis>s

We examined the effects of 2,4-epibrassinolide (EBR) application on photosynthesis, antioxidant enzyme activity, and Rubisco activase (RCA) gene expression in wheat (Triticum aestivum L.) seedlings under a combination of drought and heat stress. The net photosynthetic rates (P_n) of wheat seedlings decreased significantly, the photosynthetic capability was inhibited, and the activities of superoxide (SOD), peroxidase (POD), catalase (CAT), and RCA as well as the initial and total activity of Rubisco declined under the combined stress. These decreases and inhibitory effects were significantly ameliorated by exogenous EBR application. Three subunits (45–46, 41–42, and 38–39 kDa) of RCA were observed in wheat seedlings. The abundances of the 38–39 kDa and 41–42 kDa subunits were significantly lower in plants subjected to stressful conditions than in unstressed plants. Interestingly, a marked increase in 45–46 kDa RCA was observed under heat or heat combined with drought stress. The abundance of 38–39 kDa RCA in seedlings exposed to heat, drought, or their combination was significantly enhanced by EBR pretreatment, which paralleled the changes in initial Rubisco activity and P_n, but was not consistent with observed mRNA abundance. These results indicated that the larger subunit of RCA (45–46 kDa), which is more thermostable and increased in response to moderate heat stress, and the smaller isoform (38–39 kDa) of RCA may play important roles in maintaining the photosynthetic capability by EBR under stress conditions.     

Independent and combined abiotic stresses affect the physiology and expression patterns of DREB genes differently in stress‐susceptible and resistant genotypes of banana

In tropics, combined stresses of drought and heat often reduce crop productivity in plants like Musa acuminata L. We compared responses of two contrasting banana genotypes, namely the drought‐sensitive Grand Nain (GN; AAA genome) and drought tolerant Hill banana (HB; AAB genome) to individual drought, heat and their combination under controlled and field conditions. Drought and combined drought and heat treatments caused greater reduction in leaf relative water content and greater increase in ion leakage and H₂O₂ content in GN plants, especially in early stages, while the responses were more pronounced in HB at later stages. A combination of drought and heat increased the severity of responses. Real‐time expression patterns of the A‐1 and A‐2 group DEHYDRATION‐RESPONSIVE ELEMENT BINDING (DREB) genes revealed greater changes in expression in leaves of HB plants for both the individual stresses under controlled conditions compared to GN plants. A combination of heat and drought, however, activated most DREB genes in GN but surprisingly suppressed their expression in HB in controlled and field conditions. Its response seems correlated to a better stomatal control over transpiration in HB and a DREB‐independent pathway for the more severe combined stresses unlike in GN. Most of the DREB genes had abscisic acid (ABA)‐responsive elements in their promoters and were also activated by ABA suggesting at least partial dependence on ABA. This study provides valuable information on physiological and molecular responses of the two genotypes to individual and combined drought and heat stresses.