Comparative protein profiles of <Emphasis Type="Italic">Butea superba</Emphasis> tubers under seasonal changes

Seasonal changes are major factors affecting environmental conditions which induce multiple stresses in plants, leading to changes in protein relative abundance in the complex cellular plant metabolic pathways. Proteomics was applied to study variations in proteome composition of Butea. superba tubers during winter, summer and rainy season throughout the year using two-dimensional polyacrylamide gel electrophoresis coupled with a nanoflow liquid chromatography coupled to electrospray ionization quadrupole-time-of-flight tandem mass spectrometry. A total of 191 protein spots were identified and also classified into 12 functional groups. The majority of these were mainly involved in carbohydrate and energy metabolism (30.37 %) and defense and stress (18.32 %). The results exhibited the highest numbers of identified proteins in winter-harvested samples. Forty-five differential proteins were found in different seasons, involving important metabolic pathways. Further analysis indicated that changes in the protein levels were due mainly to temperature stress during summer and to water stress during winter, which affected cellular structure, photosynthesis, signal transduction and homeostasis, amino-acid biosynthesis, protein destination and storage, protein biosynthesis and stimulated defense and stress mechanisms involving glycolytic enzymes and relative oxygen species catabolizing enzymes. The proteins with differential relative abundances might induce an altered physiological status within plant tubers for survival. The work provided new insights into the better understanding of the molecular basis of plant proteomes and stress tolerance mechanisms, especially during seasonal changes. The finding suggested proteins that might potentially be used as protein markers in differing seasons in other plants and aid in selecting B. superba tubers with the most suitable medicinal properties in the future.     

Proteomic analysis of cold stress-responsive proteins in <Emphasis Type="Italic">Thellungiella</Emphasis> rosette leaves

Low temperature is one of the most severe environmental factors that impair plant growth and agricultural production. To investigate how Thellungiella halophila, an Arabidopsis-like extremophile, adapts to cold stress, a comparative proteomic approach based on two-dimensional electrophoresis was adopted to identify proteins that changed in abundance in Thellungiella rosette leaves during short term (6 h, 2 and 5 days) and long term (24 days) exposure to cold stress. Sixty-six protein spots exhibited significant change at least at one time point and maximal cold stress induced-proteome change was found in long-term cold stress group while the minimal change was found in 6-h cold treatment group. Fifty protein spots were identified by mass spectrometry analysis. The identified proteins mainly participate in photosynthesis, RNA metabolism, defense response, energy pathway, protein synthesis, folding and degradation, cell wall and cytoskeleton and signal transduction. These proteins might work cooperatively to establish a new homeostasis under cold stress. Nearly half of the identified cold-responsive proteins were associated with various aspects of chloroplast physiology suggesting that the cold stress tolerance of T. halophila is achieved, at least partly, by regulation of chloroplast function. All protein spots involved in RNA metabolism, defense response, protein synthesis, folding and degradation were found to be upregulated markedly by cold treatment, indicating enhanced RNA metabolism, defense and protein metabolism may play crucial roles in cold tolerance mechanism in T. halophila.     

Comparative proteomic analysis of <Emphasis Type="Italic">Phalaenopsis</Emphasis> leaves in the vegetative and flowering phase

Phalaenopsis, an epiphytic crassulacean acid metabolism (CAM) plant, requires moderate variations of day/night temperatures for flowering. In this study, changes in chlorophyll content, chlorophyll fluorescence, sugar components, titratable acidity and soluble protein content in Phalaenopsis leaves during flowering were observed. Comparative proteomic analysis of Phalaenopsis leaves in the vegetative and flowering phase was performed for the first time using iTRAQ (isobaric tags for relative and absolute quantification). A total of 126 proteins were differentially expressed in Phalaenopsis leaves. Analysis of potential functions revealed that the major categories of predicted function of the up-regulated proteins were protein destination (27 %), photosynthesis (15.9 %), primary metabolism (14.3 %) and defense (12.7 %) in the flowering phase, while the major categories of predicted function of the down-regulated proteins were protein destination (33.3 %), primary metabolism (20.6 %), transportation (14.3 %) and signal transduction (11.1 %). Proteome profile analysis indicated that the proteome changes were consistent with changes in sugar and protein metabolites. Some novel proteins were differentially expressed, most of which were identified as signaling proteins, including 14-3-3 proteins, fibrillin, rapid alkalinization factors (RALF), the Ras-related protein RABB1c, calreticulin and calmodulin. Histone, importin alpha, multidrug resistance proteins and the ABC transporters were also differentially expressed. These results provide insights into the mechanisms that regulate flowering in complex flowering plants.     

Quantitative changes in protein expression of cadmium-exposed poplar plants

Cadmium (Cd) pollution is a worldwide major concern having, among others, deleterious effects on plants. In the present work, the effects of a 20 microM Cd exposure in hydroponics culture during 14 days were evaluated in young poplar leaves. Proteins were analysed by 2-D DIGE, followed by MALDI-TOF-TOF identification. Additionally, growth and other physiological parameters were monitored during the experiment. Treated plants exhibited an inhibition of growth and visual symptoms appeared after 7 days. A significant accumulation of Cd in all organs was recorded by ICP-MS analysis. A number of changes in the expression of proteins with various functions were identified; in particular a decreased abundance of oxidative stress regulating proteins, whereas pathogenesis-related proteins showed a drastic increase in abundance. Furthermore, a large number of proteins involved in carbon metabolism showed a decrease in abundance, while proteins involved in remobilizing carbon from other energy sources were upregulated. In conclusion, the negative effect of Cd could be explained by a deleterious effect on protein expression from the primary carbon metabolism and from the oxidative stress response mechanism. Accumulation of Cd in stems of poplar, coupled with a low impact of Cd on physiological parameters, promotes the use of poplar trees for phytoremediation purposes.     

Leaf proteomic analysis in cassava (<Emphasis Type="Italic">Manihot esculenta</Emphasis>, Crantz) during plant development,from planting of stem cutting to storage root formation

Tuberization in cassava (Manihot esculenta Crantz) occurs simultaneously with plant development, suggesting competition of photoassimilate partitioning between the shoot and the root organs. In potato, which is the most widely studied tuber crop, there is ample evidence suggesting that metabolism and regulatory processes in leaf may have an impact on tuber formation. To search for leaf proteins putatively involved in regulating tuber generation and/or development in cassava, comparative proteomic approaches have been applied to monitor differentially expressed leaf proteins during root transition from fibrous to tuberous. Stringent cross comparison and statistical analysis between two groups with different plant ages using Student’s t test with 95% significance level revealed a number of protein spots whose abundance were significantly altered (P < 0.05) during week 4 to week 8 of growth. Of these, 39 spots were successfully identified by ion trap LC–MS/MS. The proteins span various functional categories from antioxidant and defense, carbohydrate metabolism, cyanogenesis, energy metabolism, miscellaneous and unknown proteins. Results suggested possible metabolic switches in the leaf that may trigger/regulate storage root initiation and growth. This study provides a basis for further functional characterization of differentially expressed leaf proteins, which can help understand how biochemical processes in cassava leaves may be involved in storage root development.     

Frerea indica,a stem succulent CAM plant with deciduous C3 leaves

Summary During the winter in greenhouse culture, Frerea indica(Asclepiadaceae) is a leafless stem succulent resembling the other members of the Stapelieae subfamily. However, in spring it produces leaves which persist during the summer period. CO₂ exchange measurements were carried out with Frerea indica at its different seasonal states of development. The leafless stems in winter as well as the defoliated ones in summer show all characteristics of Crassulacean acid metabolism. The leaves, on the other hand, fix CO₂ with high rates according to the C₃ pathway; no diurnal malate fluctuations are detectable. This feature can be interpreted as an ecological adaptation where a seasonal plant dimorphism enables a functional change in the prevailing mode of CO₂ metabolism. In the leafless state, which probably represents the appearance of the plant during the dry period in nature, it is saving water by pursuing CAM. The season with enough water available can be used for high photosynthetic gain by C₃ leaves.     

Plasmatic protein values in captive adult Iberian red deer stags (<Emphasis Type="Italic">Cervus elaphus hispanicus</Emphasis>)

The aim of this study was to assess the time trend of plasmatic proteins in red deer stags. Blood samples were taken monthly from 17 male red deer for 22 months. Total plasmatic determination and protein electrophoresis were performed. Plasmatic proteins showed minimum values during spring and summer and a maximum at the peak of the mating period. Total globulins, β and γ, followed a pattern similar to that observed for total proteins, whereas α1 and α2 globulins showed no seasonal variations. Albumin showed higher values in early spring and summer and lower values at the beginning of autumn, coinciding with the mating season. These seasonal changes in plasmatic proteins should be taken into account when assessing blood protein analysis results.     

Comparative proteomic analysis of male and female plants in Jojoba (<Emphasis Type="Italic">Simmondsia chinensis</Emphasis>) leaves revealed changes in proteins involved in photosynthesis,metabolism, energy,and biotic and abiotic stresses

Jojoba is a dioecious shrub with female and male flowers in separated individuals. The plant native to North and Central American deserts, it’s cultivated in many other places worldwide for its valuable liquid wax. The male-biased ratio in cultivated jojoba affects the yield. To develop protein molecular markers for early gender differentiation, comparative proteomic study been conducted on male and female leaves. Using gel-based proteomic, 45 proteins were identified representing 19 different proteins with 18 known functions. The identified proteins were involved in photosynthesis, energy, metabolism and the respond to biotic and abiotic stress. Ribulose-1,5-bisphosphate carboxylase (Rubisco) and ATP synthase were the most abundant proteins in both male and female of jojoba leaves, both were upregulated in male compared to female. Both proteins have the potential to serve as protein biomarkers for early differentiation between male and female in jojoba plant. These results could help in better understanding the molecular mechanism of gender differentiation in jojoba.     

Seasonal changes in the dietary compositions of rusa deer in Pangandaran Nature Reserve,West Java,Indonesia

Information regarding the food habits of tropical cervids is limited. We studied the food habits of rusa deer (Rusa timorensis), a tropical cervid in the Pangandaran Nature Reserve, West Java, Indonesia. The faecal composition of the rusa deer collected over a year was analysed using microscopy of plant fragments. The results showed marked seasonal changes: in the rainy season, the rusa deer mainly fed on Cynodon dactylon, a lawn-like grass, which accounted for approximately 50% of the faecal composition. During the dry season, C. dactylon in the faeces decreased to approximately 20%, while the composition of fruits increased, suggesting that food conditions for the deer were better in this season. This pattern differs from that of temperate cervids which can feed on abundant plant leaves during summer but are forced to consume low-quality foods such as bark and dead leaves during winter.     

Seasonal Changes in the Photosynthetic Rate in Apple Trees : A Comparison between Fruiting and Nonfruiting Trees

Seasonal changes in photosynthesis of apple trees (Malus domestica Borkh.) were monitored to examine the effect of source-sink interactions on photosynthesis and photorespiration. Elevated photosynthetic rates were observed during two periods of the growing season and correlated with the fruiting process. The first period of increased photosynthetic rates was during the bloom period, when spur leaves on flowering shoots exhibited up to 25% higher photosynthetic rates than vegetative spur leaves on a leaf area basis. CO₂ assimilation rates were also higher in fruiting trees than nonfruiting trees during the period of rapid fruit growth from July to September. Photorespiration, dark respiration, leaf resistance, and transpiration exhibited no seasonal changes which correlated to the presence or absence of fruit. These data represent the first comprehensive examination of the effects of flowering/fruit formation on photosynthesis and photorespiration in perennial plants.     

Seasonal dynamics of photosynthesis and total carbon gain in bearing and nonbearing pistachio (<Emphasis Type="Italic">Pistacia vera</Emphasis> L.) shoots

Seasonal changes in leaf gas exchange, assimilation response to light and leaf area were monitored in bearing and nonbearing pistachio shoots. Shoot bearing status did not directly affect leaf photosynthetic rate. However, photosynthetic light-response curves strongly varied during the season demonstrating the dominant effect of the tree’s seasonal phenology on assimilation. Early in the season low photosynthetic rates were associated with high rates of dark respiration indicating limited photosynthesis in the young leaves. As leaves matured, dark respiration decreased and assimilation reached maximum values. Photosynthetic efficiency was strongly reduced late in the season due to leaf age and senescence. Fruit load precipitated an early leaf senescence and drop that resulted in a 53% decrease in leaf area in bearing vs. nonbearing shoots, strongly decreasing the seasonal photosynthetic performance of bearing shoots. Bearing shoots produced a 26% lower seasonal carbon gain compared to nonbearing shoots.     

Comparative analyses of the proteomes of leaves and flowers at various stages of development reveal organ‐specific functional differentiation of proteins in soybean

The functional differentiation of protein networks in individual organs and tissues of soybean at various developmental stages was investigated by proteomic approach. Protein extraction by Mg/NP‐40 buffer followed by alkaline phenol‐based method was optimized for proteomic analysis. Proteome analyses of leaves at various developmental stages showed 26 differentially expressed proteins, wherein proteins in translocon at the outer/inner envelope membrane of chloroplast protein‐transport machineries increased significantly at the first trifoliate. Immunoblot analysis showed chaperonin‐60 expressed abundantly in young leaves, whereas HSP 70 and ATP‐synthase β were constitutively expressed in all tissues. The net photosynthesis rate and chlorophyll content showed an age‐dependent correlation in leaves. These results suggest that proteins involved in carbon assimilation, folding and assembly, and energy may work synchronously and show a linear correlation to photosynthesis at developmental stages of leaves. Comparison of flower bud and flower proteome reveals 29 differentially expressed proteins, wherein proteins involved in mitochondrial protein transport and assembly, secondary metabolism, and pollen‐tube growth were up‐regulated during flower development. Together, these results suggest that during developmental stages, each type of tissue is associated with a specific group of proteins; wherein proteins involved in energy, sugar metabolism, and folding, assembly, and destination may play pivotal roles in the maturation process of each organ or tissue.     

<Emphasis Type="SmallCaps">P</Emphasis>lastic responses to temporal variation in moisture availability: consequences for water use efficiency and plant performance

The ability to appropriately modify physiological and morphological traits in response to temporal variation should increase fitness. We used recombinant hybrid plants generated by crossing taxa in the Piriqueta caroliniana complex to assess the effects of individual leaf traits and trait plasticities on growth in a temporally variable environment. Recombinant hybrids were used to provide a wide range of trait expression and to allow an assessment of the independent effects of individual traits across a range of genetic backgrounds. Hybrid genotypes were replicated through vegetative propagation and planted in common gardens at Archbold Biological Station in Venus, Florida, where they were monitored for growth, leaf morphological characters, and integrated water use efficiency (WUE) (C isotope ratio; δ¹³C) for two successive seasons. Under wet conditions only leaf area had significant effects on plant growth, but as conditions became drier, growth rates were greatest in plants with narrow leaves and higher trichome densities. Plants with higher WUE exhibited increased growth during the dry season but not during the wet season. WUE during the dry season was increased for plants with smaller, narrower leaves that had higher trichome densities and increased reflectance. Examination of alternative path models revealed that during the dry season leaf traits had significant effects on plant growth only through their direct effects on WUE, as estimated from δ¹³C. Over the entire growing season, plants with a greater ability to produce smaller and narrower leaves with higher trichome densities in response to reduced water availability had the greatest growth rate. These findings suggest that plants making appropriate changes to leaf morphology as conditions became dry had increased WUE, and that the ability to adjust leaf phenotypes in response to environmental variation is a mechanism by which plants increase fitness.     

Proteome analysis of leaves from the resurrection plant <Emphasis Type="Italic">Boea hygrometrica</Emphasis> in response to dehydration and rehydration

Resurrection plants differ from other species in their unique ability to survive desiccation. In order to understand the mechanisms of desiccation tolerance, proteome studies were carried out using leaves of the resurrection plant Boea hygrometrica to reveal proteins that were differentially expressed in response to changes in relative water content. This opportunity was afforded by the rare ability of excised B. hygrometrica leaves to survive and resume metabolism following desiccation in a manner similar to intact plants. From a total of 223 proteins that were reproducibly detected and analyzed, 35% showed increased abundance in dehydrated leaves, 5% were induced in rehydrated leaves and 60% showed decreased or unchanged abundance in dehydrated and rehydrated leaves. Since the induction kinetics fall into clearly defined patterns, we suggest that programmed regulation of protein expression triggered by changes of water status. Fourteen dehydration responsive proteins were analyzed by mass spectrometry. Eight proteins were classified as playing a role in reactive oxygen species scavenging, photosynthesis and energy metabolism. In agreement with these findings, glutathione content and polyphenol oxidase activity were found to increase upon dehydration and rapid recovery of photosynthesis was observed.     

Geographical variation and population structure in the White-rumped Sandpiper Calidris fuscicollis as shown by morphology, mitochondrial DNA and carbon isotope ratios

Several plant-herbivore hypotheses are based on the assumption that plants cannot simultaneously allocate resources to growth and defence. We studied seasonal patterns in allocation to growth and putatively defensive compounds by monitoring several chemical and physical traits in the leaves of mountain birch from early June (budburst) to late September (leaf senescence). We found significant seasonal changes in all measured characteristics, both in terms of concentrations (mg g^-1) and amounts (mg leaf^-1). Changes were very rapid in the spring, slow in the middle of the season, and there was another period of fast changes in the senescing leaves. Co-occurring changes in physical leaf traits and concentrations of several compounds indicated a seasonal decline in foliage suitability for herbivores. Concentrations of protein and free amino acids declined through the growing season whereas individual sugars showed variable seasonal patterns. The seasonal trends of phenolic groups differed drastically: concentrations of soluble proanthocyanidins increased through the season, whereas cell wall-bound proanthocyanidins, gallotannins and flavonoid glycosides declined after an initial increase in young leaves. We failed to find proof that the seasonal accumulation of phenolics would have been seriously compromised by leaf or shoot growth, as assumed by the growth/differentiation balance hypothesis and the protein competition model hypothesis. On the contrary, there was a steady increase in the total amount of phenolics per leaf even during the most active leaf growth.     

Cold Acclimation in Genetically Related (Sibling) Deciduous and Evergreen Peach (Prunus persica [L.] Batsch): I. Seasonal Changes in Cold Hardiness and Polypeptides of Bark and Xylem Tissues

Seasonal patterns of proteins and of cold hardiness were characterized in bark and xylem tissues of genetically related (sibling) deciduous and evergreen peach (Prunus persica [L.] Batsch). In contrast with deciduous trees, which entered endodormancy and abscised leaves in the fall, evergreen trees retained their leaves and exhibited shoot elongation under favorable environmental conditions. A successive increase in the cold hardiness of bark and xylem was observed during the fall in both genotypes. This was followed by a subsequent decrease from midwinter to spring. Xylem tissue in both genotypes exhibited deep supercooling and a significant correlation (r = 0.99) between the midpoint of the low-temperature exotherm and the subzero temperature at which 50% injury occurred (assessed by electrolyte leakage) was noted. The maximum hardiness level attained in deciduous trees was more than twofold that of evergreens. Seasonal pattern of proteins from bark and xylem of the sibling genotypes was characterized by one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Among other qualitative and quantitative changes, accumulation of a 19-kilodalton polypeptide in the bark of both genotypes was observed during fall followed by a decrease in spring. This polypeptide accumulated to higher levels in the deciduous peach compared with the evergreen. Additionally, a 16-kilodalton protein exhibited the same pattern in deciduous trees but not in the evergreen trees. Both the 19- and a 16-kilodalton bark proteins conform to the criteria of a bark storage protein. The relationship of seasonal changes in protein to cold hardiness and dormancy in these genetically related peach genotypes is discussed.