Xiong Li

Comparative physiological and proteomic analyses of poplar (Populus yunnanensis) plantlets exposed to high temperature and drought.

Plantlets of Populus yunnanensis Dode were examined in a greenhouse for 48 h to analyze their physiological and proteomic responses to sustained heat, drought, and combined heat and drought. Compared with the application of a single stress, simultaneous treatment with both stresses damaged the plantlets more heavily. The plantlets experienced two apparent response stages under sustained heat and drought. During the first stage, malondialdehyde and reactive oxygen species (ROS) contents were induced by heat, but many protective substances, including antioxidant enzymes, proline, abscisic acid (ABA), dehydrin, and small heat shock proteins (sHSPs), were also stimulated. The plants thus actively defended themselves against stress and exhibited few pathological morphological features, most likely because a new cellular homeostasis was established through the collaborative operation of physiological and proteomic responses. During the second stage, ROS homeostasis was overwhelmed by substantial ROS production and a sharp decline in antioxidant enzyme activities, while the synthesis of some protective elements, such as proline and ABA, was suppressed. As a result, photosynthetic levels in P. yunnanensis decreased sharply and buds began to die, despite continued accumulation of sHSPs and dehydrin. This study supplies important information about the effects of extreme abiotic environments on woody plants.

Physiological and Proteomics Analyses Reveal the Mechanism of Eichhornia crassipes Tolerance to High-Concentration Cadmium Stress Compared with Pistia stratiotes

Cadmium (Cd) pollution is an environmental problem worldwide. Phytoremediation is a convenient method of removing Cd from both soil and water, but its efficiency is still low, especially in aquatic environments. Scientists have been trying to improve the ability of plants to absorb and accumulate Cd based on interactions between plants and Cd, especially the mechanism by which plants resist Cd. Eichhornia crassipes and Pistia stratiotes are aquatic plants commonly used in the phytoremediation of heavy metals. In the present study, we conducted physiological and biochemical analyses to compare the resistance of these two species to Cd stress at 100 mg/L. E. crassipes showed stronger resistance and was therefore used for subsequent comparative proteomics to explore the potential mechanism of E. crassipes tolerance to Cd stress at the protein level. The expression patterns of proteins in different functional categories revealed that the physiological activities and metabolic processes of E. crassipes were affected by exposure to Cd stress. However, when some proteins related to these processes were negatively inhibited, some analogous proteins were induced to compensate for the corresponding functions. As a result, E. crassipes could maintain more stable physiological parameters than P. stratiotes. Many stress-resistance substances and proteins, such as proline and heat shock proteins (HSPs) and post translational modifications, were found to be involved in the protection and repair of functional proteins. In addition, antioxidant enzymes played important roles in ROS detoxification. These findings will facilitate further understanding of the potential mechanism of plant response to Cd stress at the protein level.

Physiological, biochemical and proteomics analysis reveals the adaptation strategies of the alpine plant Potentilla saundersiana at altitude gradient of the Northwestern Tibetan Plateau

UNLABELLED This study presents an analysis of leave and rood morphology, biochemical and proteomics approach as adaptation strategies of the alpine plant Potentilla saundersiana in an altitude gradient. Several plant physiological parameter, including root and leaf architecture, leaf photosynthesis capacity, specific leaf area (SLA) and leaf nitrogen concentration, histology and microscopy, anthocyanin and proline contents, antioxidant enzyme activity assay, in-gel enzyme activity staining, H2O2 and O2(-) content, immunoblotting, auxin and strigolactone content and proteomics analysis were evaluated at five different altitudes. P. saundersiana modulated the root architecture and leaf phenotype to enhance adaptation to alpine environmental stress through mechanisms that involved hormone synthesis and signal transduction, particularly the cross-talk between auxin and strigolactone. Furthermore, an increase of antioxidant proteins and primary metabolites as a response to the alpine environment in P. saundersiana was observed. Proteins associated with the epigenetic regulation of DNA stability and post-translational protein degradation was also involved in this process. Based on these findings, P. saundersiana uses multiple strategies to adapt to the high-altitude environment of the Alpine region. BIOLOGICAL SIGNIFICANCE The alpine environment, which is characterized by sharp temperature shifts, high levels of ultraviolet radiation exposure, and low oxygen content, limits plant growth and distribution. Alpine plants have evolved strategies to survive the extremely harsh conditions prevailing at high altitudes; however, the underlying mechanisms remain poorly understood. The alpine plant Potentilla saundersiana is widespread in the Northwestern Tibetan Plateau. Here we adopted a comparative proteomics approach to investigate the mechanisms by which P. saundersiana withstands the alpine environment by examining plants located at five different altitudes. We detected and functionally characterized 118 proteins spots with variable abundance. Proteins involved in antioxidant activity, primary metabolites, epigenetic regulation, and protein post-translational modification play important roles in conferring tolerance to alpine environments. Furthermore, our results indicate that P. saundersiana modulates the root architecture and leaf phenotype to enhance adaptation to alpine environmental stress. These results provide novel insight into the multiple strategies underlying P. saundersiana adaptation to the high-altitude environment of the Northwestern Tibetan Plateau.

Molecular cloning and characterization of a novel SK3-type dehydrin gene from Stipa purpurea

Stipa purpurea, an endemic forage of the grass family in the Tibetan Plateau, is highly resistant to drought. Dehydrins (DHNs) are stress proteins involved in plant protective reactions against environmental stress. In this study, the full-length DHN open reading frame (ORF) cloned from S. purpurea, named SpDHN1, was 816 nucleotides length and encoded a protein of 271 amino acids. Phylogenetic and sequence characterization analysis revealed that the DHN gene was an SK3-type DHNs. Subcellular localization analysis indicated that SpDHN1 was localized in the cytoplasm and the plasma membrane. SpDHN1 function analysis provided new evidence to support the antioxidation of SpDHN1 in plant responses drought stress. Ectopic expression of SpDHN1 in Arabidopsis thaliana plants showed more resistance to drought stress than the wild-type, indicating that SpDHN1 may be a potential candidate gene for genetic improvement of crops to improve stress tolerance.

Physiological and Proteomic Adaptation of the Alpine Grass Stipa purpurea to a Drought Gradient

Stipa purpurea, an endemic forage species on the Tibetan Plateau, is highly resistant to cold and drought, but the mechanisms underlying its responses to drought stress remain elusive. An understanding of such mechanisms may be useful for developing cultivars that are adaptable to water deficit. In this study, we analyzed the physiological and proteomic responses of S. purpurea under increasing drought stress. Seedlings of S. purpurea were subjected to a drought gradient in a controlled experiment, and proteins showing changes in abundance under these conditions were identified by two-dimensional electrophoresis followed by mass spectrometry analysis. A western blotting analysis was conducted to confirm the increased abundance of a heat-shock protein, NCED2, and a dehydrin in S. purpurea seedlings under drought conditions. We detected carbonylated proteins to identify oxidation-sensitive proteins in S. purpurea seedlings, and found that ribulose-1, 5-bisphosphate carboxylase oxygenase (RuBisCO) was one of the oxidation-sensitive proteins under drought. Together, these results indicated drought stress might inhibit photosynthesis in S. purpurea by oxidizing RuBisCO, but the plants were able to maintain photosynthetic efficiency by a compensatory upregulation of unoxidized RuBisCO and other photosynthesis-related proteins. Further analyses confirmed that increased abundance of antioxidant enzymes could balance the redox status of the plants to mitigate drought-induced oxidative damage.

Transcriptome analysis reveals diversified adaptation of Stipa purpurea along a drought gradient on the Tibetan Plateau

Natural selection drives species adaptations to biotic and abiotic stresses. Species distributed along a moisture gradient, such as Stipa purpurea, a dominant grass in alpine arid and semi-arid meadows on the Tibetan Plateau, provide an opportunity to evaluate the effects of long-term adaptation to differing degrees of drought stress on gene expression. However, the genetic basis of this divergence remains largely unknown. Next-generation sequencing technologies have provided important genome-wide insights on the evolution of organisms for which genomic information is lacking. To understand how S. purpurea responds to drought stress, we selected five populations distributed along the degressive rainfall line on the northwestern Tibetan Plateau that currently present evolutionary acclimation to localized drought pressure at the physiological and biochemical levels and compared their transcriptome responses. In addition, we performed de novo assembly of the S. purpurea transcriptome using short read sequencing technology and successfully assembled 84,298 unigenes from approximately 51 million sequencing reads. We quantified gene expression level to compare their transcriptome responses using mRNA-Seq and identified differentially expressed transcripts that are involved in primary and secondary plant metabolism, plant hormone synthesis, defense responses, and cell wall synthesis. Furthermore, physiological and biochemical evidence supports that abscisic acid (ABA) accumulation and cell wall strengthening derived from the differential transcripts contribute to the tolerance of S. purpurea to drought stress. The mechanisms by which S. purpurea adapts to drought stress provide new insight into how plants ecologically adapt and evolve.

A novel Ap2/ERF transcription factor from Stipa purpurea leads to enhanced drought tolerance in Arabidopsis thaliana.

Key messageSpERF1 acts as a positive regulator, contributing to drought stress tolerance inA. thalianathrough activating DRE/CRT elements in the promoters of abiotic stress-responsive genes.AbstractStipa purpurea is an endemic perennial grass species in alpine arid and semi-arid meadows on the Qinghai-Xizang Plateau, which is highly tolerant against drought and cold. ERF transcription factors are known to regulate gene expression under abiotic and biotic treatments. Herein, we isolated a full-length ERF gene CDS from S. purpurea named SpERF1, which was induced by drought, cold, and jasmonic acid stresses. Subcellular localization revealed that SpERF1 is a nuclear protein, consistent with its roles as a transcription factor. Overexpression of SpERF1 enhanced drought tolerance of transgenic Arabidopsis thaliana via the activation of DRE/CRT elements in the promoters of abiotic stress-responsive genes. Furthermore, increased accumulation of proline indicated that SpERF1 might be involved in proline synthesis in the transgenic lines, allowing them to have a better buffering capacity and membrane protection under drought stress. This study indicated that SpERF1 might be an attractive target in the genetic engineering for improving stress tolerance in other crops. Moreover, SpERF1 protein function analysis increased our understanding of S. purpurea’s ability to adapt to the adverse conditions of the Qinghai-Xizang Plateau.

Comparative Physiological and Proteomic Analysis Reveals the Leaf Response to Cadmium-Induced Stress in Poplar (Populus yunnanensis).

Excess amounts of heavy metals are important environmental pollutants with significant ecological and nutritional effects. Cdmium (Cd) is of particular concern because of its widespread occurrence and high toxicity. We conducted physiological and proteomic analyses to improve our understanding of the responses of Populus yunnanensis to Cd stress. The plantlets experienced two apparent stages in their response to Cd stress. During the first stage, transiently induced defense-response molecules, photosynthesis- and energy-associated proteins, antioxidant enzymes and heat shock proteins (HSPs) accumulated to enhance protein stability and establish a new cellular homeostasis. This activity explains why plant photosynthetic capability during this period barely changed. During the second stage, a decline of ribulose-1, 5-bisphosphate carboxylase (RuBisCO) and HSP levels led to imbalance of the plant photosynthetic system. Additionally, the expression of Mitogen-activated protein kinase 3 (MPK3), Mitogen-activated protein kinase 6 (MPK6) and a homeobox-leucine zipper protein was higher in the second stage. Higher expression of caffeoyl-CoA O-methyltransferase (CCoAOMT) may regulate plant cell wall synthesis for greater Cd storage. These genes may be candidates for further research and use in genetic manipulation of poplar tolerance to Cd stress.

Activation of Secondary Cell Wall Biosynthesis by miR319‐targeted TCP4 transcription factor

Summary The overexpression of miR319 in plants results in delayed senescence, and high levels of miR319‐targeted TCP4 transcription factor cause premature onset of this process. However, the underlying mechanisms of this pathway remain elusive. Here, we found that miR319 overexpression results in a decrease in TCP4 abundance and secondary cell wall formation in the stem. Conversely, constitutive expression of miR319‐resistant TCP4 promotes secondary cell wall formation, indicating that miR319‐mediated TCP4 controls secondary cell wall formation during development. Further analysis revealed that TCP4 might directly bind the promoter of VND7 to activate its expression, which triggers the expression of a VND7 transcriptional network associated with secondary cell wall biosynthesis and programmed cell death and accelerates vessel formation. In addition, the development process gradually increased TCP4 expression. These results suggest that miR319 and its target TCP4 can act as switches that turn on secondary cell wall synthesis and programmed cell death.

Expression of Stipa purpurea SpCIPK26 in Arabidopsis thaliana Enhances Salt and Drought Tolerance and Regulates Abscisic Acid Signaling

Stipa purpurea (S. purpurea) is the dominant plant species in the alpine steppe of the Qinghai-Tibet Plateau, China. It is highly resistant to cold and drought conditions. However, the underlying mechanisms regulating the stress tolerance are unknown. In this study, a CIPK gene from S. purpurea (SpCIPK26) was isolated. The SpCIPK26 coding region consisted of 1392 bp that encoded 464 amino acids. The protein has a highly conserved catalytic structure and regulatory domain. The expression of SpCIPK26 was induced by drought and salt stress. SpCIPK26 overexpression in Arabidopsis thaliana (A. thaliana) plants provided increased tolerance to drought and salt stress in an abscisic acid (ABA)-dependent manner. Compared with wild-type A. thaliana plants, SpCIPK26-overexpressing plants had higher survival rates, water potentials, and photosynthetic efficiency (Fv/Fm), as well as lower levels of reactive oxygen species (ROS) following exposure to drought and salt stress. Gene expression analyses indicated stress-inducible genes (RD29A, RD29B, and ABF2) and a ROS-scavenger gene (CAT1) were upregulated in SpCIPK26-overexpressing plants after stress treatments. All of these marker genes are associated with ABA-responsive cis-acting elements. Additionally, the similarities in the gene expression patterns following ABA, mannitol, and NaCl treatments suggest SpCIPK26 has an important role during plant responses to drought and salt stress and in regulating ABA signaling.