Proteomic changes in response to low-light stress during cotton fiber elongation

Numerous studies have illustrated that low light is one of the major abiotic stresses limiting cotton (Gossypium hirsutum L.) fiber length, but studies addressing molecular mechanisms contributing to reduced fiber growth under low light are lacking. To investigate the molecular mechanisms of cotton fiber elongation in response to low light, an experiment of low light caused by shading was conducted with cotton cultivar NuCOTN 33B. The results showed that low light resulted in shorter fiber length. Proteomic analysis of four developmental stages (5, 10, 15 and 20 days post-anthesis) showed that 49 proteins were significantly responsive to low light. 39 differentially expressed proteins that included some known as well as some novel low-light stress-responsive proteins were identified. These differentially expressed proteins were involved in signal transduction, carbohydrate/energy metabolism, cell wall component synthesis, protein metabolism, cytoskeleton, nitrogen metabolism and stress responses. The results also showed that the decrease in fiber length might be because the levels of signal-related protein (phospholipase D), cytoskeletal proteins (two annexins isoforms), cell wall component-related proteins (sucrose synthase, UDP-d-glucuronic acid 4-epimerase and rhamnose synthase), carbohydrate metabolism-proteins (phosphofructokinase, dihydrolipoamide dehydrogenase, vacuolar H⁺-ATPase catalytic subunit, malate dehydrogenase and isocitrate dehydrogenase), and stress-related proteins (peroxisomal catalase, short chain alcohol dehydrogenase) were decreased under low light.     

Protein expression changes during cotton fiber elongation in response to low temperature stress

Low temperature stress is one of the major abiotic stresses limiting the formation of cotton (Gossypium hirsutum L.) fiber qualities, especially fiber length. To investigate the molecular adaptation mechanisms of cotton fiber elongation to low temperature stress, two cotton cultivars, Kemian 1 (low temperature-tolerant) and Sumian 15 (low temperature-sensitive), were planted in the field at two sowing dates (25 April and 10 June). The two sowing dates resulted in different growing conditions and the main environmental difference between them was temperature, particularly the mean daily minimum temperature (MDTmin). When the sowing date was delayed, the MDTmin decreased from 26.9 °C (25 April) to 20.6 °C (10 June). Low temperature stress (MDTmin of 20.6 °C) shortened the fiber length significantly in two cultivars, but the decreased extent was larger in Sumian 15 than that in Kemian 1. Proteomic analysis of three developmental stages (10, 15 and 20 days post-anthesis [DPA]) showed that 37 spots changed significantly (p < 0.05) in abundance under low temperature stress and they were identified using mass spectrometry. These proteins were involved in malate metabolism, soluble sugar metabolism, cell wall loosening, cellulose synthesis, cytoskeleton, cellular response, and redox homeostasis. The results suggest that the enhancement of osmoticum maintenance, cell wall loosening, cell wall components biosynthesis, and cytoskeleton homeostasis plays important roles in the tolerance of cotton fibers to low temperature stress. Moreover, low levels of PEPCase, expansin, and ethylene signaling proteins may potentially lead to the low temperature sensitivity of Sumian 15 at the proteomic level.     

A high‐confidence reference dataset of differentially expressed proteins in elongating cotton fiber cells

In our previous study, we used a comparative proteomic approach based on 2DE to profile dynamic proteomes of cotton fibers and found 235 protein spots differentially expressed during the elongation process ranging from 5 to 25 days post‐anthesis. Of them, only 106 differentially expressed proteins (DEPs) were identified by MS due to database limitations at the time. In the present work, we successfully identified the remaining 129 DEPs from the same experimental system using high‐resolution MS with an updated database. Bioinformatic analysis revealed that proteins involved in carbohydrate and protein metabolism, transport, and redox homeostasis are the most abundant, and glycolysis was found to be the most significantly regulated process during fiber elongation. Our high‐confidence reference dataset, composed of 235 DEPs, provides a valuable resource for future studies on the molecular mechanism of cotton fiber elongation.     

iTRAQ‐based quantitative proteomic analysis gives insight into sexually different metabolic processes of poplars under nitrogen and phosphorus deficiencies

Male and female poplars (Populus cathayana Rehd.) respond differently to nitrogen (N) and phosphorus (P) deficiencies. In this study, an iTRAQ‐based quantitative proteomic analysis was performed. N and P deficiencies caused 189 and 144 proteins to change in abundance in males and 244 and 464 in females, respectively. Compared to N‐ and P‐deficient males, both N‐ and P‐deficient females showed a wider range of changes in proteins that are involved in amino acid, carbohydrate and protein metabolism, and the sexual differences were significant. When comparing the effects of N‐ and P‐deficiencies, N‐deficient females expressed more changes in proteins that are involved in stress responses and gene expression regulation, while P‐deficient females showed more changes in proteins that are involved in energy and lipid metabolism, stress responses and gene expression regulation. The quantitative RT‐PCR analysis of stress‐related proteins showed that males have a better expression correlation between mRNA and protein levels than do females. This study shows that P. cathayana females are more sensitive and have more rapid metabolic mechanisms when responding to N and P deficiencies than do males, and P deficiency has a wider range of effects on females than does N deficiency.     

Salt tolerance is unrelated to carbohydrate metabolism in cowpea cultivars

Under salinity stress, plants commonly accumulate carbohydrates for osmotic adjustment to balance the excess accumulated ions and to protect biomolecules. We selected two cowpea cultivars with contrasting response to salinity, Pitiúba (salt-tolerant) and TVu (salt-sensitive), to investigate whether the salt tolerance could be associated with changes in carbohydrate accumulation and metabolism in leaves and roots during a long-term experiment. Two salt treatments (0 and 75 mM NaCl) were applied to 10-day-old plants grown in nutrient solution for 24 days. Despite some changes in carbohydrate accumulation and carbohydrate metabolism enzymes induced by salt stress, no consistent alterations in carbohydrates could be found in leaves or roots in this study. Therefore, we suggest that tolerance to salt stress is largely unrelated to carbohydrate accumulation in cowpea.     

Protein expression changes during cotton fiber elongation in response to drought stress and recovery

An investigation to better understand the molecular mechanism of cotton (Gossypium hirsutum L.) fiber elongation in response to drought stress and recovery was conducted using a comparative proteomics analysis. Cotton plants (cv. NuCOTN 33B) were subjected to water deprivation for 10 days followed by a recovery period (with watering) of 5 days. The temporal changes in total proteins in cotton fibers were examined using 2DE. The results revealed that 163 proteins are significantly drought responsive. MS analysis led to the identification of 132 differentially expressed proteins that include some known as well as some novel drought‐responsive proteins. These drought responsive fiber proteins in NuCOTN 33B are associated with a variety of cellular functions, i.e. signal transduction, protein processing, redox homeostasis, cell wall modification, metabolisms of carbon, energy, lipid, lignin, and flavonoid. The results suggest that the enhancement of the perception of drought stress, a new balance of the metabolism of the biosynthesis of cell wall components and cytoskeleton homeostasis plays an important role in the response of cotton fibers to drought stress. Overall, the current study provides an overview of the molecular mechanism of drought response in cotton fiber cells.     

氮素对不同开花期棉铃纤维比强度形成的生理基础的影响

于2005年在江苏南京(长江流域下游棉区)和徐州(黄河流域黄淮棉区)棉田设置不同氮素水平(零氮：0 kg N·hm^-2,适氮：240 kg N·hm^-2,高氮：480 kg N·hm^-2)试验,研究氮素对不同开花期棉铃(伏前桃、伏桃和秋桃)纤维比强度形成生理基础的影响.结果表明：与适氮处理相比,零氮处理显著降低了棉铃对位叶氮浓度,增加了C/N,影响程度随开花期的推迟而加大,导致伏桃、秋桃对位叶制造和运输光合产物的能力在棉铃发育中后期大幅度下降,棉纤维的相对生长速率以及纤维发育关键酶蔗糖合成酶和β-1,3-葡聚糖酶活性降低,纤维素快速累积持续期缩短,纤维比强度显著降低;高氮处理显著增加了棉铃对位叶氮浓度,降低了C/N,影响程度随开花期的推迟而降低,其降低了伏前桃、伏桃发育过程中光合产物向纤维分配的比例、棉铃发育前中期的纤维发育关键酶活性及纤维素累积速率,导致其纤维比强度亦显著降低.综合分析认为,适宜的施氮量可以协调棉花的“源库”关系,有利于促进不同开花期棉铃高纤维比强度的形成.与适氮处理相比,零氮处理的伏前桃、伏桃和秋桃纤维比强度分别降低了1.8%、5.8%和13.0%,高氮处理则分别降低了8.2%、7.4%和-2.4%.     

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...     

Water-deficit stress effects on pistil biochemistry and leaf physiology in cotton (Gossypium hirsutum,L.)

Flowering in cotton (Gossypium hirsutum L.) is a sensitive stage to water-deficit stress, but the effects on metabolism are not well understood. The objective of this study was to monitor gas exchange responses of cotton plants under conditions of limited water supply and evaluate the effects on the carbohydrate concentrations and glutathione reductase levels in the cotton flower. Growth chamber experiments were conducted in 2008 and 2009, with normal day/night conditions of 32/24 °C and optimum quantities of Hoagland's nutrient solution applied until flowering. Treatments were imposed at flowering and consisted of control (Control), where optimum quantities of water were applied, and water stress (WS) where 50% of optimum quantity of water was supplied. Water-deficit stress resulted in a significant decrease in leaf stomatal conductance. Leaf photosynthetic and respiration rates were similarly decreased compared to the control. Ovary and style water potential of water-stressed leaves were significantly higher compared to the water potential of water stressed leaves, indicating that cotton flowers are fairly resistant to changes in the water status of the plant. However, carbohydrate concentrations of water-stressed pistils (ovary and style) were significantly increased compared to the control and a similar pattern was observed in the levels of glutathione reductase of water-stressed pistils. In conclusion, water-deficit stress during flowering resulted in significant decreases in leaf gas exchange functions as well as leaf water potential. Cotton pistils appeared to be less sensitive since they were able to maintain water potential similar to the control under limited water supply and increase glutathione reductase levels. However, pistil carbohydrate metabolism was significantly affected resulting in accumulation of both hexose and sucrose indicating a perturbation in sucrose cleaving and hexose utilizing enzymes that could potentially have as a consequence a decrease in fertilization and seed set efficiency.     

Effect of Pre-Stressing on the Acid-Stress Response in Bifidobacterium Revealed Using Proteomic and Physiological Approaches

Weak acid resistance limits the application of Bifidobacteria as a probiotic in food. The acid tolerance response (ATR), caused by pre-stressing cells at a sublethal pH, could improve the acid resistance of Bifidobacteria to subsequent acid stress. In this study, we used Bifidobacterium longum sub. longum BBMN68 to investigate the effect of the ATR on the acid stress response (ASR), and compared the difference between the ATR and the ASR by analyzing the two-dimensional-PAGE protein profiles and performing physiological tests. The results revealed that a greater abundance of proteins involved in carbohydrate metabolism and protein protection was present after the ASR than after the ATR in Bifidobacterium. Pre-stressing cells increased the abundance of proteins involved in energy production, amino acid metabolism, and peptidoglycan synthesis during the ASR of Bifidobacterium. Moreover, after the ASR, the content of ATP, NH₃, thiols, and peptidoglycan, the activity of H⁺-ATPase, and the maintenance of the intracellular pH in the pre-stressed Bifidobacterium cells was significantly higher than in the uninduced cells. These results provide the first explanation as to why the resistance of Bifidobacterium to acid stress improved after pre-stressing.     

Overexpression of <Emphasis Type="Italic">ScALDH21</Emphasis> gene in cotton improves drought tolerance and growth in greenhouse and field conditions

Aldehyde dehydrogenase (ALDH) is essential for scavenging redundant aldehydes when plants are exposed to stress. The aim of the present study was to validate the ectopic expression of the ScALDH21 gene, which is isolated from Syntrichia caninervis, an extremely drought-tolerant moss, to improve drought tolerance in cotton (Gossypium hirsutum L.). In our study, the ScALDH21-transformed cotton was identified via PCR, RT-PCR, and DNA gel blotting, and the growth and physiological characteristics related to drought tolerance were compared between the transgenic cotton (TC) and non-transgenic cotton (NT) grown in a greenhouse and in field conditions. The results indicated that TC accumulated approximately 11.8–304 % more proline than did NT under drought stress, and produced a lower concentration of lipid peroxidation-derived reactive aldehydes and had a higher peroxidase activity under oxidative stress. Moreover, TC showed reduced loss of the net photosynthetic rate compared with NT. Under field conditions, TC showed greater plant height, larger bolls, and greater cotton fiber yield than NT, but no significant difference in fiber quality between TC and NT following different water-withholding treatments. These results suggest that overexpression of ScALDH21 can greatly improve the drought tolerance of cotton without reduction in yield and fiber quality.     

施氮方式对转基因棉花Bt蛋白含量及产量的影响

为研究氮肥运筹对棉花叶片、棉蕾和棉铃不同器官中Bt蛋白含量的影响,2009—2010年,以抗虫杂交棉中棉所72为试验材料,在大田条件下进行了不同基肥：花铃肥：盖顶肥施氮比例（分别为0:0.4:0.6、0.2:0.4:0.4、0.4:0.4:0.2、0.6:0.4:0）的试验。结果表明,施氮方式对棉花不同器官中Bt蛋白含量有明显影响。总体表现为随着氮肥前移,棉花幼嫩器官中Bt蛋白含量呈明显增加的趋势,而老熟器官中Bt蛋白含量呈明显降低的趋势。施氮方式对棉花幼嫩器官中Bt蛋白含量的影响比老熟器官明显,尤其是对幼嫩叶片Bt蛋白含量的影响大于幼小的棉蕾和棉铃器官。抗虫棉采用基肥：花铃肥：盖顶肥为0.4:0.4:0.2的施氮方式,总体能提高前中期棉花器官Bt蛋白的含量,有利于提高其抗虫性能;减少后期棉花器官Bt蛋白的含量,减轻对环境的压力;而且比其余3种施氮方式的籽棉产量和皮棉产量分别增加4.15%—11.24％、3.73%—12.01%。     

Involvement of ubiquitin-proteasome system in icariin-induced cardiomyocyte differentiation of embryonic stem cells using two-dimensional gel electrophoresis

Icariin has been shown to significantly facilitate the differentiation of embryonic stem (ES) cells into cardiomyocytes in vitro. However, the mechanism underlying the icariin-induced cardiomyocyte differentiation is still not fully understood. In the present study, 52 differentially displayed proteins selected from two-dimensional electrophoresis gels were identified by MALDI-TOF mass spectrometry analysis. More than half of proteins could be assigned to six main categories: (1) protein synthesis, metabolism, processing and degradation, (2) stress response, (3) cytoskeleton proteins, (4) energy metabolism, (5) carbohydrate metabolism/transport, and (6) RNA/other nucleic acids metabolisms and transport, nuclear proteins. MALDI-TOF/MS showed that icariin treatment resulted in the induction of five ubiquitin-proteasome system (UPS)-related proteins, such as ubiquitin carboxy-terminal hydrolase L1 (UCH-L1), ubiquitin-conjugating enzyme E2N, proteasome 26S, proteasome subunit-alpha type 6, and proteasome subunit-alpha type 2 in the differentiated cardiomyocytes. These results implied that UPS might play an important role in the control of cardiomyocyte differentiation. Epoxomicin (a proteasome inhibitor) significantly reduced the cardiomyocyte differentiation rate of ES cells and proteasome activities, as well as inhibited NF-κB translocation into the nucleus, which were evidently reversed by presence of icariin. Meanwhile, icariin could significantly reverse the reduction of four proteins (proteasome subunit-alpha type 6, proteasome subunit-alpha type 2, UCH-L1, and ubiquitin-conjugating enzyme E2N) expressions owing to application of epoxomicin. These suggest UPS could be a means by which icariin may regulate expressions of key proteins that control cardiomyocyte differentiation. Taken together, these results indicated that UPS played an important role in ES cell differentiate into cardiomyocytes induced by icariin.     

Effect of shading on yield,fiber quality and physiological characteristics of cotton subtending leaves on different fruiting positions

Little is known regarding to impact of simulated shading conditions on cotton yield and fiber quality at different fruiting positions. In this 2-year study, our field experiments investigated the effects of shading percentage on the cotton yield, fiber properties, photosynthesis, and carbohydrate concentrations in boll’s subtending leaves during various growing stages at different fruiting positions (FP). Net photosynthetic rate and effective quantum yield of PSII photochemistry decreased in response to shading on both FP1 and FP3 of the 7^th sympodial branches, respectively. Shading also reduced sucrose and starch contents of leaves at each fruiting position. Shading decreased the number and mass of cotton bolls, the fiber strength and micronaire, while the fiber length increased at both fruiting positions. Our results suggested that shading resulted in the reduction of the cotton yield and fiber quality, which are mainly associated with the changes in boll number and alteration of photosynthesis and carbohydrate concentrations during the boll development.