ABSTRACT: Jasmonic acid (JA) and methyl jasmonate (MeJA) regulate plant development, resistance to stress, and insect attack by inducing specific gene expression. However, little is known about the mechanism of plant defense against herbivore attack at a protein level. Using a high-resolution 2-DE gel, we identified 60 MeJA-responsive proteins and measured protein expression level changes. Among these 62 proteins, 43 proteins levels were increased while 11 proteins were decreased. We also found eight proteins uniquely expressed in response to MeJA treatment. The proteins identified in this study have important biological functions including photosynthesis and energy related proteins (38.4%), protein folding, degradation and regulated proteins (15.0%), stress and defense regulated proteins (11.7%), and redox-responsive proteins (8.3%). We found MeJA could not only induce plant defense mechanisms to insects, it also enhanced toxic protein production that potentially can be used for bio-control of Asian corn borer.
Project description:Spermatophyte pollen tubes and root hairs have been used as single-cell-type model systems toward understanding the molecular processes underlying polar growth of plant cells. Horsetail (Equisetum arvense L.) is a perennial herb species in Equisetopsida, which creates separately growing spring and summer stems in its life cycle. The mature chlorophyllous spores produced from spring stems can germinate without dormancy. Here we report the cellular features and protein expression patterns in five stages of horsetail spore germination (mature spores, rehydrated spores, double-celled spores, germinated spores, and spores with protonemal cells). Using 2-DE combined with mass spectrometry, 80 proteins were found to be differentially expressed upon spore germination. Among them, proteins involved in photosynthesis, protein turnover, and energy supply were over-represented. Thirteen proteins appeared as isoforms on the gels, indicating the potential importance of post-translational modification. In addition, the dynamic changes of ascorbate peroxidase, peroxiredoxin, and dehydroascorbate reductase implied that reactive oxygen species homeostasis is critical in regulating cell division and tip-growth. The time course of germination and diverse expression patterns of proteins in photosynthesis, energy supply, lipid and amino acid metabolism indicated that heterotrophic and autotrophic metabolism were necessary in light-dependent germination of the spores. Twenty-six proteins were involved in protein synthesis, folding, and degradation, indicating that protein turnover is vital to spore germination and rhizoid tip-growth. Furthermore, the altered abundance of small G protein Ran1, 14-3-3 protein, actin, and Caffeoyl-CoA O-methyltransferase revealed that signaling transduction, vesicle trafficking, cytoskeleton dynamics, and cell wall modulation were critical to cell division and polar growth. These findings provide up-to-date evidences for understanding fern spore asymmetric division and rhizoid polar growth.
Project description:Cryoinjury and protein changes are a consequence of cryopreservation and may have a negative impact on sperm quality regarding motility, viability and fertilizing ability. However, potential proteomic changes in rabbit semen throughout the cryopreservation process have never been investigated previously. The aim of the present study was to compare the whole proteome of fresh and cryopreserved rabbit extracellular fluid of semen. Comparative analysis and identification of proteins were performed using 2-dimensional difference in-gel electrophoresis (2D-DIGE) coupled with matrix-assisted laser desorption/ionization mass (MALDI TOF/TOF) spectrometry. We found that 28 proteins differed in abundance between fresh and frozen extracellular fluid.
Project description:Serine protease is considered a relative upstream regulator in the physiological processes including male reproduction. Transmembrane serine protease 12 (TMPRSS12) has been shown to regulate sperm motility and uterotubal junction migration in mice, but its role in the testis remains unknown.In this study, we confirmed the important role of TMPRSS12 in spermatogenesis. To further investigate the mechanism of TMPRSS12 in male reproduction, we used 2-D electrophoresis combined with Matrix-assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry(MALDI-TOF/TOF) to construct differential protein expression profiles of testes in Tmprss12-/- and WT mice. A total of 64 spots with 2-fold or more differential expression between two groups were identified using MALDI-TOF/TOF. As a result, 54 unique proteins were identified successfully, including 39 proteins with decreased expression and 15 proteins with increased expression in Tmprss12-/- mice.
Project description:Cryoinjury and protein changes are a consequence of cryopreservation and may have a negative impact on sperm quality regarding motility, viability and fertilizing ability. However, potential proteomic changes in rabbit semen throughout the cryopreservation process have never been investigated previously. The aim of the present study was to compare the whole proteome of fresh and cryopreserved rabbit spermatozoa. Comparative analysis and identification of proteins were performed using 2-dimensional difference in-gel electrophoresis (2D-DIGE) coupled with matrix-assisted laser desorption/ionization mass (MALDI TOF/TOF) spectrometry. We found that 108 proteins differed in abundance in spermatozoa between fresh and frozen semen. Most of these proteins was involved in pathways related to energy metabolism and protein quality control under stress conditions, reproductive processes and mechanisms of cell death/survival regulation, resulting in a significant decrease of motility and viability of post-thawing rabbit sperm and its potential fertilizing ability.
Project description:Gall formation on the belowground parts of plants infected with Plasmodiophora brassicae is the result of extensive host cellular reprogramming. The development of these structures is a consequence of increased cell proliferation followed by massive enlargement of cells colonised with the pathogen. Drastic changes in cellular growth patterns create local deformities in the roots and hypocotyl giving rise to mechanical tensions within the tissue of these organs. Host cell wall extensibility and recomposition accompanies growth of the gall, influences pathogen spread and also pathogen life cycle progression. Demethylation of pectin within the extracellular matrix may play an important role in P. brassicae-driven hypertrophy of host underground organs. Through proteomic analysis of the cell wall we identified proteins accumulating in the galls developing on the underground parts of Arabidopsis thaliana plants infected with P. brassicae. One of the key proteins identified was the pectin methylesterase PME18; we further characterised its expression and conducted functional and anatomic studies in the knock-out mutant and used Raman spectroscopy to study the status of pectin in P. brassicae infected galls. We found that late stages of gall formation are accompanied with increased levels of Pectin Methylesterase 18 (PME18). We have also shown, that the massive enlargement of cells colonised with P. brassicae coincides with decreases in pectin methylation. In pme18-1 knock-out mutants P. brassicae could still induce demethylation; however, the galls in this line were smaller and cellular expansion was less pronounced. Alteration in pectin demethylation in the host resulted in changes in pathogen distribution and slowed down disease progression. To conclude, P. brassicae driven host organ hypertrophy observed during clubroot disease is accompanied by pectin demethylation in the extracellular matrix. The pathogen hijacks endogenous host mechanisms involved in cell wall loosening to create an optimal cellular environment for completion of its life cycle and eventual release of resting spores facilitated by degradation of demethylated pectin polymers.
Project description:Lung cancer is responsible for the most cancer-related mortality worldwide and the mechanism of its development is poorly understood. Proteomics has become a powerful tool offering vital knowledge related to cancer development. Using a two-dimensional difference gel electrophoresis (2D-DIGE) approach, we sought to compare tissue samples from non-small-cell lung cancer (NSCLC) patients, exactly squamous cell carcinoma (SCC), taken from the tumor center and tumor margin and control (adjacent non-tumor) tissues. We found and 21 spots representing 20 proteins differentiating center and margin. We found 111 differentially expressed protein spots representing 95 proteins; 84, 7, and 20 spots differed between the control and center and margin, control and center, and control and margin, respectively. Nine significant canonical pathways were identified, including hypoxia-inducible factor-1α signaling, thyroid hormone biosynthesis, and phagosome maturation. Several pathways related to disease and function were identified as related to cell death and survival, and several related to cancer, organismal injury, and abnormalities. Proteins differentiating the tumor center and tumor margin were linked to cancer invasion and progression, including cell migration, adhesion and invasion, cytoskeletal structure, protein folding, anaerobic metabolism, tumor angiogenesis, epithelialmesenchymal transition, epithelial adherens junctions, and inflammatory responses.
Project description:The liquid storage of turkey semen without the loss of fertilizing ability is of practical interest to poultry industry. However, fertility rates from liquid stored turkey semen decline within a few hours. A clear cause of the decline in spermatozoa quality is unknown. Therefore, the purpose of the present study was to monitor dynamic of proteomic changes of spermatozoa during liquid storage at three time points (2, 24 and 48h) by 2-dimensional difference in-gel electrophoresis (2D-DIGE) coupled with a matrix-assisted laser desorption/ionization mass (MALDI TOF/TOF) spectrometry. A total of 57 protein spots were differentially expressed in fresh and stored spermatozoa; 42 spots were more- and 15 were less- abundant in spermatozoa after 48 h of semen storage. Most of the proteins that changed in response to liquid storage were related to i) flagellum dependent cell motility, ii) energy derivation by oxidation of organic compounds and iii) introduction of fertilization, suggesting complexity of the processes leading to decreases of stored semen quality.
Project description:Lung cancer is responsible for the most cancer-related mortality worldwide and the mechanism of its development is poorly understood. Proteomics has become a powerful tool offering vital knowledge related to cancer development. Using a two-dimensional difference gel electrophoresis (2D-DIGE) approach, we sought to compare tissue samples from non-small-cell lung cancer (NSCLC) patients, exactly adenocarcinoma (ADC), taken from the tumor center and tumor margin and control (adjacent non-tumor) tissues . We found 22 differentially abundant spots representing 17 proteins differentiating center and margin. Sixty eight proteins; represented by 79 spots :40, 20, and 19 spots differed between the control and center and margin, control and center, and control and margin, respectively. Twenty six significant canonical pathways were identified, including Rho signaling pathways, a semaphorin neuronal repulsive signaling pathway, and epithelial adherens junction signaling. Proteins differentiating the tumor center and tumor margin were linked to cancer invasion and progression, including cell migration, adhesion and invasion, cytoskeletal structure, protein folding, anaerobic metabolism, tumor angiogenesis, EMC transition, epithelial adherens junctions, and inflammatory responses.
Project description:During Agrobacterium rhizogenes–plant interaction, the rolB gene is transferred into the plant genome and is stably inherited in the plant’s offspring. Among the numerous effects of rolB on plant metabolism, including the activation of secondary metabolism, its effect on plant defense systems has not been sufficiently studied. Here, we performed a proteomic analysis of rolB-expressing Arabidopsis plants with a particular focus on defense proteins. In rolB plants, a reduced amount of scaffold proteins RACK1A, RACK1B, and RACK1C, known as receptors for activated C-kinase 1, was found. Proteomic analysis showed that rolB could potentially suppress the plant immune system by suppressing the RNA-binding proteins GRP7, CP29B, and CP31B, similar to the action of type III bacterial effectors. At the same time, rolB plants induce massive biosynthesis of protective proteins VSP1, VSP2, and PR4, which are markers of the activated jasmonate pathway. Moreover, rolB plants activate all components of the PYK10 defense complex of the endoplasmic reticulum involved in the metabolism of glucosinolates. We hypothesized that various defense systems activated by rolB are aimed at protecting the host plant from competing phytopathogens in order to create an effective ecological niche for A. rhizogenes. A RolB → RACK1A signaling module has been proposed that can exert most of the rolB-mediated effects on plant physiology and biochemistry.
Project description:ECB larvae remain damaging pests of conventiational maize production. To better understand the physiological and molecular changes occuring in stem tissue during insect attack we explored the biochemical and transcriptional profiles in these tissues. We used microarrays to detail the transcriptional changes associated with ECB attack and considered these in the context of biochemical markers of plant defense and total protein levels. Maize internode tissue was selected from untreated control plants and those which 5th instar ECB larvae had bored into for 48 h for RNA extraction and hybridization on Affymetrix microarrays.