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Switchgrass plants were grown in a Sandwich tube system to induce gradual drought stress by withholding watering. After 29 days, the leaf photosynthetic rate decreased significantly, compared to the control plants which were watered regularly. The drought-treated plants recovered to the same leaf water content after three days of re-watering. The root tip (1cm basal fragment, designated as RT1 hereafter) and the elongation/maturation zone (the next upper 1 cm tissue, designated as RT2 hereafter) tissues were collected at the 29th day of drought stress treatment, (named SDT for severe drought treated), after one (D1W) and three days (D3W) of re-watering. The tandem mass tags mass spectrometry-based quantitative proteomics analysis was performed to identify the proteomes, and drought-induced differentially accumulated proteins (DAPs). From RT1 tissues, 6156, 7687, and 7699 proteins were quantified, and 296, 535, and 384 DAPs were identified in the SDT, D1W, and D3W samples, respectively. From RT2 tissues, 7382, 7255, and 6883 proteins were quantified, and 393, 587, and 321 proteins DAPs were identified in the SDT, D1W, and D3W samples. Between RT1 and RT2 tissues, very few DAPs overlapped at SDT, but the number of such proteins increased during the recovery phase. A large number of hydrophilic proteins and stress-responsive proteins were induced during SDT and remained at a higher level during the recovery stages. A large number of DAPs in RT1 tissues maintained the same expression pattern throughout drought treatment and the recovery phases. The DAPs in RT1 tissues were classified in cell proliferation, mitotic cell division, and chromatin modification, and those in RT2 were placed in cell wall remodeling and cell expansion processes. This study provided information pertaining to root zone-specific proteome changes during drought and recover phases, which will allow us to select proteins (genes) as better defined targets for developing drought tolerant plants. The mass spectrometry proteomics data are available via ProteomeXchange with identifier PXD017441.

In this paper, we report on aluminum (Al)-induced root proteomic changes in switchgrass. After growth in a hydroponic culture system supplemented with 400 μM of Al, plants began to show signs of physiological stress such as a reduction in photosynthetic rate. At this time, the basal 2-cm long root tips were harvested and divided into two segments, each of 1-cm in length, for protein extraction. Al-induced changes in proteomes were identified using tandem mass tags mass spectrometry (TMT-MS)-based quantitative proteomics analysis. A total of 216 proteins (approximately 3.6% of total proteins) showed significant differences between non-Al treated control and treated groups with significant fold change (twice the standard deviation; FDR adjusted p-value < 0.05). The apical root tip tissues expressed more dramatic proteome changes (164 significantly changed proteins; 3.9% of total proteins quantified) compared to the elongation/maturation zones (52 significantly changed proteins, 1.1% of total proteins quantified). Significantly changed proteins from the apical 1-cm root apex tissues were clustered into 25 biological pathways; proteins involved in the cell cycle (rotamase FKBP 1 isoforms, and CDC48 protein) were all at a reduced abundance level compared to the non-treated control group. In the root elongation/maturation zone tissues, the identified proteins were placed into 18 pathways, among which proteins involved in secondary metabolism (lignin biosynthesis) were identified. Several STRING protein interaction networks were developed for these Al-induced significantly changed proteins. This study has identified a large number of Al-responsive proteins, including transcription factors, which will be used for exploring new Al tolerance genes and mechanisms. Data are available via ProteomeXchange with identifiers PXD008882 and PXD009125.

The tomato (Solanum lycopersicum) ripening process from mature green (MG) to turning and then to red stages is accompanied by the occurrences of physiological and biochemical reactions, which ultimately result in the formation of the flavor, color and texture of ripe fruits. The two trivalent metal ions Al3+ and La3+ are known to induce different levels of phytotoxicity in suppressing root growth. This paper aims to understand the impacts of these two metal ions on tomato fruit proteomes. Tomato ‘Micro-Tom’ plants were grown in a hydroponic culture system supplemented with 50 μM aluminum sulfate (Al2 (SO4)3.18H2O) for Al3+ or La2(SO4)3 for La3+. Quantitative proteomics analysis, using isobaric tags for relative and absolute quantitation, were performed for fruits at MG, turning and red stages. Results show that in MG tomatoes, proteins involved in protein biosynthesis, photosynthesis and primary carbohydrate metabolisms were at a significantly lower level in Al-treated compared to La-treated plants. For the turning and red tomatoes, only a few proteins of significant differences between the two metal treatments were identified. Results from this study indicate that compared to La3+, Al3+ had a greater influence on the basic biological activities in green tomatoes, but such an impact became indistinguishable as tomatoes matured into the late ripening stages.

This study aimed to identify the aluminum (Al)-induced proteomes in tomato (Solanum lycopersicum, "Micro-Tom") after long-term exposure to the stress factor. Plants were treated in Magnavaca's solution (pH 4.5) supplemented with 7.5 μM Al(3+) ion activity over a 4 month period beginning at the emergence of flower buds and ending when the lower mature leaves started to turn yellow. Proteomes were identified using a 8-plex isobaric tags for relative and absolute quantification (iTRAQ) labeling strategy followed by a two-dimensional (high- and low-pH) chromatographic separation and final generation of tandem mass spectrometry (MS/MS) spectra of tryptic peptides on an LTQ-Orbitrap Elite mass spectrometer. Principal component analysis revealed that the Al-treatment had induced systemic alterations in the proteomes from roots and leaves but not seed tissues. The significantly changed root proteins were shown to have putative functions in Al(3+) ion uptake and transportation, root development, and a multitude of other cellular processes. Changes in the leaf proteome indicate that the light reaction centers of photosynthetic machinery are the primary targets of Al-induced stress. Embryo and seed-coat tissues derived from Al-treated plants were enriched with stress proteins. The biological processes involving these Al-induced proteins concur with the physiological and morphological changes, such as the disturbance of mineral homeostasis (higher contents of Al, P, and Fe and reduced contents of S, Zn, and Mn in Al-treated compared to nontreated plants) in roots and smaller sizes of roots and the whole plants. More importantly, the identified significant proteins might represent a molecular mechanism for plants to develop toward establishing the Al tolerance and adaptation mechanism over a long period of stress treatment.

Aluminum (Al) toxicity is a major constraint to plant growth and crop yield in acid soils. Tomato cultivars are especially susceptible to excessive Al3+ accumulated in the root zone. In this study, tomato plants were grown in a hydroponic culture system supplemented with 50 µM AlK(SO₄)₂. Seeds harvested from Al-treated plants contained a significantly higher Al content than those grown in the control hydroponic solution. In this study, these Al-enriched tomato seeds (harvested from Al-treated tomato plants) were germinated in 50 µM AlK(SO₄)₂ solution in a homopiperazine-1,4-bis(2-ethanesulfonic acid) buffer (pH 4.0), and the control solution which contained the buffer only. Proteomes of radicles were analyzed quantitatively by mass spectrometry employing isobaric tags for relative and absolute quantitation (iTRAQ®). The proteins identified were assigned to molecular functional groups and cellular metabolic pathways using MapMan. Among the proteins whose abundance levels changed significantly were: a number of transcription factors; proteins regulating gene silencing and programmed cell death; proteins in primary and secondary signaling pathways, including phytohormone signaling and proteins for enhancing tolerance to abiotic and biotic stress. Among the metabolic pathways, enzymes in glycolysis and fermentation and sucrolytic pathways were repressed. Secondary metabolic pathways including the mevalonate pathway and lignin biosynthesis were induced. Biological reactions in mitochondria seem to be induced due to an increase in the abundance level of mitochondrial ribosomes and enzymes in the TCA cycle, electron transport chains and ATP synthesis.

Switchgrass (Panicum virgatum) is a perennial crop producing deep roots and thus highly tolerant to soil water deficit conditions. However, seedling establishment in the field is very susceptible to prolonged and periodic drought stress. In this study, a “sandwich” system simulating a gradual water deletion process was developed. Switchgrass seedlings were subjected to a 20-day gradual drought treatment process when soil water tension was increased to 0.05 MPa (moderate drought stress) and leaf physiological properties had expressed significant alteration. Drought-induced changes in leaf proteomes were identified using the isobaric tags for relative and absolute quantitation (iTRAQ) labeling method followed by nano-scale liquid chromatography mass spectrometry (nano-LC-MS/MS) analysis. Additionally, total leaf proteins were processed using a combinatorial library of peptide ligands to enrich for lower abundance proteins. Both total proteins and those enriched samples were analyzed to increase the coverage of the quantitative proteomics analysis. A total of 7006 leaf proteins were identified, and 257 (4% of the leaf proteome) expressed a significant difference (p < 0.05, fold change 1.7) from the non-treated control to drought-treated conditions. These proteins are involved in the regulation of transcription and translation, cell division, cell wall modification, phyto-hormone metabolism and signaling transduction pathways, and metabolic pathways of carbohydrates, amino acids, and fatty acids. A scheme of abscisic acid (ABA)-biosynthesis and ABA responsive signal transduction pathway was reconstructed using these drought-induced significant proteins, showing systemic regulation at protein level to deploy the respective mechanism. Results from this study, in addition to revealing molecular responses to drought stress, provide a large number of proteins (candidate genes) that can be employed to improve switchgrass seedling growth and establishment under soil drought conditions (Data are available via ProteomeXchange with identifier PXD004675).