GpEF1A: a novel lysine methyltransferase gene from Gypsophila perfoliata L. involved in boron homeostasis.

Rapid accumulation of boron (B) leads to toxicity in plant tissues, and the narrow gap between deficiency and toxicity makes it difficult to adjust essential B levels in soil for plant productivity. Therefore, understanding different aspects of B tolerance is necessary to provide new and valid solutions to B toxicity. Gypsophila perfoliata stands out as a remarkable example of a B-tolerant plant, with a natural propensity to thrive in environments such as B mines and soils enriched with high levels of B. In this study, a yeast functional screening experiment was conducted using cDNA libraries from G. perfoliata leaf and root cells for B tolerance. Ten colonies from the leaf library grew in 80 mm boric acid, while none emerged from the root library. Analysis of isolated cDNAs showed identical sequences and a unique motif related to B tolerance. The gene GpEF1A was identified in the tolerant yeast colonies, with predicted structural features suggesting its role, and RT-qPCR indicating increased expression under B stress. A regulatory role for EF1A lysine methylation was proposed in mammalian cells and fungi because of its dynamic and inducible nature under environmental constraints. This could also be relevant for plant cells, as the high similarity of the GpEF1A gene in some salt-tolerant plants might indicate the upregulation of EF1A as a conserved way to cope with abiotic stress conditions. This report represents the first instance of involvement of GpEF1A in B tolerance, and further detailed studies are necessary to understand other components of this tolerance mechanism.

Heterophyllin B, a cyclopeptide from Pseudostellaria heterophylla, improves memory via immunomodulation and neurite regeneration in i.c.v.Aß-induced mice.

Pseudostellaria heterophylla, has historically been used as medicine food homology plant for thousand years in China. Our previous studies had indicated that daily intake of Pseudostellaria heterophylla extract enhanced cognitive memory. Herein, heterophyllin B (HET-B), a brain permeable cyclopeptide from Pseudostellaria heterophylla was determined, and the molecular mechanism underlying its memory improvement effects was investigated. Pseudostellaria heterophylla extract as well as HET-B reversed Aß25-35-induced axonal atrophy and neuronal apoptosis in cultured cortical neurons of mice. HET-B could enhance memory retrieval, modulate splenic T helper cell, and ameliorate neuroinflammation in i.c.v. Aß1-42 injected Alzheimer's disease (AD) mice. To explore the mechanism of action, network pharmacology was performed to predict protein targets and pathways of HET-B against AD. Five key targets were identified related to the effect of HET-B in AD intervention, and were clarified involved in axonal regeneration. We revealed for the first time that HET-B promoted memory retrieval through axonal regeneration and anti-neuroinflammation. This study provides a basis to research on HET-B as nutritional supplements for brain healthy.

Towards Understanding the Involvement of H+-ATPase in Programmed Cell Death of Psammosilene tunicoides after Oxalic Acid Application.

Psammosilene tunicoides is a unique perennial medicinal plant species native to the Southwestern regions of China. Its wild population is rare and endangered due to over-excessive collection and extended growth (4-5 years). This research shows that H+-ATPase activity was a key factor for oxalate-inducing programmed cell death (PCD) of P. tunicoides suspension cells. Oxalic acid (OA) is an effective abiotic elicitor that enhances a plant cell's resistance to environmental stress. However, the role of OA in this process remains to be mechanistically unveiled. The present study evaluated the role of OA-induced cell death using an inverted fluorescence microscope after staining with Evans blue, FDA, PI, and Rd123. OA-stimulated changes in K+ and Ca2+ trans-membrane flows using a patch-clamp method, together with OA modulation of H+-ATPase activity, were further examined. OA treatment increased cell death rate in a dosage-and duration-dependent manner. OA significantly decreased the mitochondria activity and damaged its electron transport chain. The OA treatment also decreased intracellular pH, while the FC increased the pH value. Simultaneously, NH4Cl caused intracellular acidification. The OA treatment independently resulted in 90% and the FC led to 25% cell death rates. Consistently, the combined treatments caused a 31% cell death rate. Furthermore, treatment with EGTA caused a similar change in intracellular pH value to the La3+ and OA application. Combined results suggest that OA-caused cell death could be attributed to intracellular acidification and the involvement of OA in the influx of extracellular Ca2+, thereby leading to membrane depolarization. Here we explore the resistance mechanism of P. tunicoides cells against various stresses endowed by OA treatment.

Target-site and non-target-site-based resistance to tribenuron-methyl in multiply-resistant Myosoton aquaticum L.

Myosoton aquaticum L., a widespread and competitive winter weed of wheat in China, has evolved resistance to many classes of herbicides. In one M. aquaticum population (AH03), collected from Anhui Province, where tribenuron-methyl and florasulam had been used to control this weed resistance to both herbicides had evolved. Compared with the sensitive population, HN03(S), the resistant (R) population, AH03, was highly resistant to tribenuron-methyl, flucarbazone-Na and pyroxsulam, moderately resistant to pyrithiobac­sodium, and florasulam, and had low resistance to diflufenican. AH03 was still controlled by imazethapyr, 2,4-D butylate, fluroxypyr-meptyl, and isoproturon. Pretreatment with the P450 inhibitor malathion reduced the GR50 value of tribenuron-methyl by 43% in the R population, and by 25% in the S population. This indicates that P450-mediated enhanced metabolism is one likely mechanism for tribenuron-methyl resistance in M. aquaticum. Glutathione-S-transferase (GST) activity could be induced by tribenuron-methyl in both the R and S populations. However, both the basal and induced GST activity of the R population was lower than that of the S population. The in vitro ALS assay confirmed that the ALS from the R plants showed a high resistance (52.93-fold) to tribenuron-methyl. ALS gene sequencing revealed a Pro197Ala substitution in the R plants. Based on the ALS gene sequence analysis, molecular markers were also developed to identify the specific Pro197Ala mutation. This population of M. aquaticum has multiple resistance and target-site (ALS Pro197Ala) and non-target-site resistance mechanisms contribute to tribenuron-methyl resistance.

Copper stress induces antioxidant responses and accumulation of sugars and phytochelatins in Antarctic Colobanthus quitensis (Kunth) Bartl.

BACKGROUND: In field, C. quitensis is subjected to many abiotic extreme environmental conditions, such as low temperatures, high UV-B, salinity and reduced water potentials, but not metal or metalloid high concentrations in soil, however, other members of Caryophyllaceae family have tolerance to high concentrations of metals, this is the case of Silene genre. In this work, we hypothesize that C. quitensis have the same mechanisms of Silene to tolerate metals, involving accumulation and induction of antioxidant systems, sugar accumulation and the induction of thiols such as phytochelatins to tolerate. RESULTS: The results showing an effective antioxidant defensive machinery involving non-enzymatic antioxidants such as phenolics, GSH and ascorbic acid, in another hand, GSH-related oligomers (phytochelatins) and sugars was induced as a defensive mechanism. CONCLUSIONS: Colobanthus quitensis exhibits certain mechanisms to tolerate copper in vitro demonstrating its plasticity to tolerate several abiotic stress conditions.

Novel Polysaccharide H-1-2 from Pseudostellaria Heterophylla Alleviates Type 2 Diabetes Mellitus.

BACKGROUND/AIMS: To investigate the potential therapeutic effect of novel polysaccharide H-1-2 from pseudostellaria heterophylla against type 2 Diabetes Mellitus (T2DM) and elucidate the underling molecular mechanisms. METHODS: Relative expression of HIF1α and Sirt1 in T2DM patients was determined via real-time PCR. The direct binding of HIF1α on Sirt1 promoter was validated by ChIP assay. The inhibitory regulation of Sirt1 by HIF1α was analyzed using luciferase reporter assay. The endogenous protein of HIF1α and Sirt1 in response to H-1-2 treatment was quantified by western blotting. The blood glucose, secreted insulin and serous lipid profiles were measured with ELISA kits. RESULTS: We consolidated that HIF1α and Sirt1 was dysregulated in T2DM patients and subjected to H-1-2 modulation. H-1-2 significantly inhibited hypoxia and up-regulated Sirt1 expression in EndoC-ßH1 cells. Accordingly, H-1-2 enhanced glucose-stimulation insulin secretion and improved blood glucose and lipid profiles in T2DM cells, and elevated the glucose and insulin tolerance simultaneously. Furthermore, we demonstrated that H-1-2 alleviated T2DM via inhibition of hypoxia and up-regulation of Sirt1 in isolated pancreatic ß-cells from T2DM rats. CONCLUSION: Our data unambiguously demonstrated H-1-2 administration alleviated T2DM by enhancing Sirt1 expression through inhibition of hypoxia.

Enhanced Herbicide Metabolism and Metabolic Resistance Genes Identified in Tribenuron-Methyl Resistant Myosoton aquaticum L.

The evolved resistance of Myosoton aquaticum L. to acetolactate synthase (ALS) inhibitors is well established, but most research has focused on target-site resistance, while nontarget-site resistance remains neglected. Here, we investigated mechanisms of the latter. The pretreatment with the P450 inhibitor malathion significantly increased the sensitivity of resistant plants to tribenuron-methyl. The rapid P450-mediated tribenuron-methyl metabolism in resistant plants was confirmed by LC-MS/MS analysis. Besides, GST activity was higher among resistant than susceptible individuals. The next transcriptome analysis generated 544,102,236 clean reads from RNA sequencing libraries. De novo assembly yielded 102,529 unigenes with an average length of 866 bp, annotated across seven databases. Digital gene expression selected 25 differentially expressed genes, further validated with qRT-PCR. Three P450 genes, two GST genes, two glucosyltransferase genes, four ABC transporter genes, and four additional contigs were constitutively up-regulated in resistant individuals. Overall, our research confirmed that enhanced herbicide metabolism drives tribenuron-methyl resistance in M. aquaticum.

Effects of indole-3-butytric acid on lead and zinc accumulations in Pseudostellaria maximowicziana.

Plant hormones can improve the phytoremediation capabilities of heavy metal hyperaccumulator plants. In this study, different doses of indole-3-butytric acid (IBA) were sprayed on the leaves of the lead (Pb) and zinc (Zn) accumulator plant Pseudostellaria maximowicziana, which was planted in Pb-Zn contaminated soil, and the effects of IBA on Pb and Zn accumulation levels in P. maximowicziana were studied. Spraying 25- and 50-mg/L IBA doses increased the stem, leaf and shoot biomasses of P. maximowicziana compared with the control, while 75- and 100-mg/L IBA doses decreased them. The 50-mg/L IBA dose increased the P. maximowicziana contents of chlorophyll a, total chlorophyll and carotenoid of compared with the control, and other doses had no significant effects or decreased these values. Spraying IBA reduced the superoxide dismutase activity of P. maximowicziana compared with the control, but improved the peroxidase and catalase activities. The 50-, 75-, and 100-mg/L IBA doses increased the Pb and Zn contents in P. maximowicziana compared with the control and also increased the amounts of Pb and Zn extracted by P. maximowicziana. Thus, 50 mg/L of IBA could promote the growth and the Pb and Zn phytoremediation capabilities of P. maximowicziana.

Copper stress induces antioxidant responses and accumulation of sugars and phytochelatins in Antarctic Colobanthus quitensis (Kunth) Bartl

BACKGROUND: In field, C. quitensis Is subjected to many abiotic extreme environmental conditions, such as low temperatures, high UV-B, salinity and reduced water potentials, but not metal or metalloid high concentrations in soil, however, other members of Caryophyllaceae family have tolerance to high concentrations of metals, this is the case of Silene genre. In this work, we hypothesize that C. quitensis have the same mechanisms of Silene to tolerate metals, involving accumulation and induction of antioxidant systems, sugar accumulation and the induction of thiols such as phytochelatins to tolerate. RESULTS: The results showing an effective antioxidant defensive machinery involving non-enzymatic antioxidants such as phenolics, GSH and ascorbic acid, in another hand, GSH-related oligomers (phytochelatins) and sugars was induced as a defensive mechanism. CONCLUSIONS: Colobanthus quitensis exhibits certain mechanisms to tolerate copper in vitro demonstrating its plasticity to tolerate several abiotic stress conditions.

Photosynthetic limitations in two Antarctic vascular plants: importance of leaf anatomical traits and Rubisco kinetic parameters.

Particular physiological traits allow the vascular plants Deschampsia antarctica Desv. and Colobanthus quitensis (Kunth) Bartl. to inhabit Antarctica. The photosynthetic performance of these species was evaluated in situ, focusing on diffusive and biochemical constraints to CO2 assimilation. Leaf gas exchange, Chl a fluorescence, leaf ultrastructure, and Rubisco catalytic properties were examined in plants growing on King George and Lagotellerie islands. In spite of the species- and population-specific effects of the measurement temperature on the main photosynthetic parameters, CO2 assimilation was highly limited by CO2 diffusion. In particular, the mesophyll conductance (gm)-estimated from both gas exchange and leaf chlorophyll fluorescence and modeled from leaf anatomy-was remarkably low, restricting CO2 diffusion and imposing the strongest constraint to CO2 acquisition. Rubisco presented a high specificity for CO2 as determined in vitro, suggesting a tight co-ordination between CO2 diffusion and leaf biochemistry that may be critical ultimately to optimize carbon balance in these species. Interestingly, both anatomical and biochemical traits resembled those described in plants from arid environments, providing a new insight into plant functional acclimation to extreme conditions. Understanding what actually limits photosynthesis in these species is important to anticipate their responses to the ongoing and predicted rapid warming in the Antarctic Peninsula.

Proteomic analysis of naturally occurring boron tolerant plant Gypsophila sphaerocephala L. in response to high boron concentration.

Gypsophila sphaerocephala is a naturally Boron (B) tolerant species that can grow around the B mining areas in Turkey, where the B concentration in soil reaches a lethal dose for plants (up to ∼8900mgkg-1 (∼140mM). While its interesting survival capacity in extremely B containing soils, any molecular research has been conducted to understand its high tolerance mechanism yet. In the present study, we have performed a proteomic analysis of this plant to understand its high tolerance towards B-stress. Seedlings of G. sphaerocephala were collected from B mining area and were adapted to greenhouse conditions. An excessive level of Boric acid (3mM)was applied to the plantlets for 24h. Total proteins were precipitated by using TCA/Acetone method. 2D-PAGE (two-dimensional polyacrylamide gel electrophoresis) analysis of the proteins was carried out. Out of 121 protein spots, 14 were differentially expressed between the control and B-exposed G. sphaerocephala roots. The peptide profile of each protein was determined by MALDI-TOF mass spectrometer after in-gel trypsin digestion. The identified proteins are involved in different mechanisms in the cell such as in antioxidant mechanism, energy metabolism, protein degradation, lipid biosynthesis and signaling pathways, indicating that G. sphaerocephala has multiple cooperating mechanisms to protect itself from high B levels. Overall, this study sheds light on to the possible regulatory switches (gene/s) controlling the B-tolerance proteins and their possible roles in plant's defense mechanism.

Production of Δ7-Sterols from In Vitro Root Cultures of Endangered Gypsophila trichotoma.

Species from the genus Gypsophila are known for their medicinal, industrial and decorative applications. G. trichotoma Wend. is an endangered plant species for the Bulgarian flora according to the Red Data Book. Δ7-Sterols, which are unusual and rare in the plant kingdom, are present in the roots of this species. In previous studies different in vitro cultures were established from aerial parts of the species. The objective of this study was to explore the possibility for production of Δ7-sterols from in vitro cultured roots of G. trichotoma. The root cultures were grown on six modified MS media and the quantity of sterols was analyzed. These findings will serve to solve the important matter of the role of nutrients on sterols biosynthesis.

Uniconazole (S-3307) strengthens the growth and cadmium accumulation of accumulator plant Malachium aquaticum.

The effects of uniconazole (S-3307) application on the growth and cadmium (Cd) accumulation of accumulator plant Malachium aquaticum (L.) Fries. were studied through a pot experiment. The application of S-3307 increased the biomass and photosynthetic pigment content of M. aquaticum in Cd-contaminated soil, and also improved the superoxide dismutase (SOD) and peroxidase (POD) activities in M. aquaticum. Application of S-3307 increased Cd content in shoots and decreased Cd content in roots of M. aquaticum, but the translocation factor (TF) of M. aquaticum increased with the increase of S-3307 concentration. For phytoextraction, the application of S-3307 increased Cd extractions by roots, shoots and whole plants of M. aquaticum, and the maxima were obtained at 75 mg L-1 S-3307, which increased by 22.07%, 37.79% and 29.07%, respectively, compared with their respective controls. Therefore, S-3307 can be used for enhancing the Cd extraction ability of M. aquaticum, and 75 mg L-1 S-3307 was the optimal dose.

Drought stress memory in the photosynthetic mechanisms of an invasive CAM species, Aptenia cordifolia.

Plants are known for their high capacity to acclimatise to fluctuating environmental conditions. A wide range of environmental conditions can lead to suboptimal physiological efficiency. However, recent studies have shown that plants can withstand repeated periods of stress. To find out how they do it, we studied photosynthetic adjustments to repeated water stress in Aptenia cordifolia: a facultative, invasive CAM species. Plants were subjected to three cycles of water deficit, and photosynthetic parameters and chloroplast antioxidants were quantified to gain an understanding of the mechanisms by which they cope with repeated stress periods. Significant modification of the photosystems' antenna and reaction centres was observed in plants subjected to previous water stress cycles, and this led to higher PSII efficiency than in plants challenged with drought for the first time. These findings underline the biological significance of stress memory and show how plants can adjust their photosynthetic apparatus to fluctuating environmental conditions and thus optimise photosynthesis and photoprotection under drought conditions.

De Novo Sequencing and Assembly Analysis of the Pseudostellaria heterophylla Transcriptome.

Pseudostellaria heterophylla (Miq.) Pax is a mild tonic herb widely cultivated in the Southern part of China. The tuberous roots of P. heterophylla accumulate high levels of secondary metabolism products of medicinal value such as saponins, flavonoids, and isoquinoline alkaloids. Despite numerous studies on the pharmacological importance and purification of these compounds in P. heterophylla, their biosynthesis is not well understood. In the present study, we used Illumina HiSeq 4000 sequencing platform to sequence the RNA from flowers, leaves, stem, root cortex and xylem tissues of P. heterophylla. We obtained 616,413,316 clean reads that we assembled into 127, 334 unique sequences with an N50 length of 951 bp. Among these unigenes, 53,184 unigenes (41.76%) were annotated in a public database and 39, 795 unigenes were assigned to 356 KEGG pathways; 23,714 unigenes (8.82%) had high homology with the genes from Beta vulgaris. We discovered 32, 095 DEGs in different tissues and performed GO and KEGG enrichment analysis. The most enriched KEGG pathway of secondary metabolism showed up-regulated expression in tuberous roots as compared with the ground parts of P. heterophylla. Moreover, we identified 72 candidate genes involved in triterpenoids saponins biosynthesis in P. heterophylla. The expression profiles of 11 candidate unigenes were analyzed by quantitative real-time PCR (RT-qPCR). Our study established a global transcriptome database of P. heterophylla for gene identification and regulation. We also identified the candidate unigenes involved in triterpenoids saponins biosynthesis. Our results provide an invaluable resource for the secondary metabolites and physiological processes in different tissues of P. heterophylla.

Interaction of Pseudostellaria heterophylla with Quorum Sensing and Quorum Quenching Bacteria Mediated by Root Exudates in a Consecutive Monoculture System.

Many plant-pathogenic bacteria are dependent on quorum sensing (QS) to evoke disease. In this study, the population of QS and quorum quenching (QQ) bacteria was analyzed in a consecutive monoculture system of Pseudostellaria heterophylla. The isolated QS strains were identified as Serratia marcescens with SwrIR-type QS system and exhibited a significant increase over the years of monoculture. Only one QQ strain was isolated from newly planted soil sample and was identified as Bacillus thuringiensis, which secreted lactonase to degrade QS signal molecules. Inoculation of S. marcescens to P. heterophylla root could rapidly cause wilt disease, which was alleviated by B. thuringiensis. Furthermore, the expression of lactonase encoded by the aiiA gene in S. marcescens resulted in reduction of its pathogenicity, implying that the toxic effect of S. marcescens on the seedlings was QS-regulated. Meanwhile, excess lactonase in S. marcescens led to reduction in antibacterial substances, exoenzymes, and swarming motility, which might contribute to pathogensis on the seedlings. Root exudates and root tuber extracts of P. heterophylla significantly promoted the growth of S. marcescens, whereas a slight increase of B. thuringiensis was observed in both samples. These results demonstrated that QS-regulated behaviors in S. marcescens mediated by root exudates played an important role in replanting diseases of P. heterophylla.

Transcriptomic analysis of Pseudostellariae Radix from different fields using RNA-seq.

Pseudostellariae Radix is an important traditional Chinese medicine (TCM), which is consumed commonly for its positive health effects. However, a lack of transcriptomic and genomic information hinders research on Pseudostellariae Radix. Here, high-throughput RNA sequencing (RNA-seq) was employed for the de novo assembly to analyze the transcriptome in Pseudostellariae Radix, finding significantly differentially expressed genes in this TCM from different fields based on RNA-seq and bioinformatic analysis. A total of 146,408,539 paired-end reads were generated and assembled into 89,857 unigenes with an average length of 862bp. All of the assembly unigenes were annotated by running BLASTx and BLASTn similarity searches on the Non-redundant nucleotide database (NT), the Non-redundant protein database (NR), Swiss-Prot, Cluster of Orthologous Groups (COG), Kyoto Encyclopedia of Genes and Genomes (KEGG), Gene Ontology (GO), and Interpro. On the basis of bioinformatic analysis and the expression profiles for Pseudostellariae Radix, 29 significantly differentially expressed genes were identified, which provides the basic information for exploring the molecular mechanisms that determine the quality of Pseudostellariae Radix from different fields. The expression levels of 29 genes were validated by real-time quantitative PCR (RT-qPCR). This is the first study to sample Pseudostellariae Radix, which provides an invaluable resource for understanding the genome of this herb.

iTRAQ-based quantitative proteomic analysis of cultivated Pseudostellaria heterophylla and its wild-type.

Pseudostellaria heterophylla is an important traditional Chinese herbal medicine with vast clinical consumption because of its positive effects. To date, changes in the metabolite composition of P. heterophylla have been well documented; however, the molecular differences between cultivated P. heterophylla and its wild-type are still unknown. The aim of this study was to investigate differences in cultivated and wild P. heterophylla. Due to the lack of a genomic database, we used high-throughput transcriptomic and proteomic technologies to identify proteins in the herb. Isobaric tags for relative and absolute quantification (iTRAQ) MS/MS were used to detect statistically significant changes between cultivated and wild P. heterophylla. We detected 3775 proteins; 332 showed differential accumulations across two different ecotypes of P. heterophylla. 71 significant differential expressions of proteins were selected based on GO annotations, KEGG, STRING analysis, and expression level. Quantitative real-time PCR analysis confirmed the authenticity and accuracy of the proteomic analysis. The results indicated that the carbohydrate and cellular amino acid metabolisms in cultivated P. heterophylla were weaker than those in its wild-type; seven important proteins were found to regulate sucrose and amino acids. This will provide the basic information for exploring the cause of differences in secondary metabolites in different ecotypes of P. heterophylla and the protein mechanism of its quality formation. BIOLOGICAL SIGNIFICANCE: This study combined a transcriptome, proteome, and metabolism approach for analyzing differentially expressed proteins of cultivated and wild P. heterophylla and established the relationship between significantly differentially expressed proteins and differential chemical components in non-model plants. The results of proteomic analysis provide the basic information for exploring the quality forming process, which will demonstrate, and provide guidance for, the study of effective constituents of P. heterophylla and its quality formation mechanism.

ECO-physiological response of S. vulgaris to CR(VI): Influence of concentration and genotype.

The objective of this work is to study the response of Silene vulgaris to a range of environmentally relevant concentrations of Cr(VI) in order to evaluate its potential use in the phytomanagement of Cr polluted sites. Cuttings of six homogenous genotypes from Madrid (Spain) have been used as plant material. The eco-physiological response of S. vulgaris to Cr(VI) changed with the genotype. The yield dose-response curve was characterized by stimulation at low doses of Cr(VI). The effects of metal concentration were quantified on root dry weight, water content and chlorophyll content, determined by SPAD index. The response was not homogeneous for all studied genotypes. At high doses of Cr(VI), plants increased micronutrient concentration in dry tissues which suggested that nutrient balance could be implicated in the alleviation of Cr toxicity. This work highlights the importance of studying the eco-physiological response of metallophytes under a range of pollutant concentrations to determine the most favorable traits to be employed in the phytomanagement process.

[iTRAQ-based quantitative proteomics of cultivated Pseudostellaria heterophylla and its wild type].

This work was designed to investigate proteins differentially expressed in cultivated Pseudostellaria heterophylla and its wild type using i TRAQ proteomics approach. The extracted proteins were digested using FASP method and identified by i TRAQ coupled with LC-MS/MS technology and then analyzed by Protein Pilot 5.0 search engine. Proteins differentially expressed were searched through comparison of relatively quantified proteins. The analysis was conducted using GO(gene ontology), KEGG and STRING. A total of 3 775 proteins were detected, among them, 3 676 proteins can be quantified, of which 127 proteins were up-regulated and 205 were down-regulated in cultivated Pseudostellaria heterophylla. We found 71 significantly differentially expressed proteins for further analysis. These proteins were classified into nine categories: heat shock proteins, transferases, oxidoreductases, lyases, isomerases, ligases, hydrolases, tubulin and translocases. The results indicated that the carbohydrate and cellular amino acids metabolism of cultivated Pseudostellaria heterophylla were weaker than its wild type and its ability of responding to stress was much stronger. GWD1, PHS1, GBE1, PGM, and BAM1 are the important proteins to regulate sucrose; met E and CYS are the key proteins that regulate amino acids in cultivated and wild Pseudostellaria heterophylla. This will provide the basic information for exploring the cause of secondary metabolites differences in different ecotype of Pseudostellaria heterophylla and the protein mechanism of its quality formation.