Project description:The experimental design was to determine the regulon of the transcriptional activator RarA. This was done by comparing the transcriptome profiles between a rarA overexpressor and the baseline control. RarA overexpression was achieved by plasmid mediated overexpression of the rarA gene and the baseline control was a vector only strain.
Project description:The effect of cell free supernatant from the antifungal strain Lactobacillus plantarum 16 on the growth of Aspergillus fumigatus Af293 was assessed. Transcriptome analysis of the genome was performed after ten minutes exposure to antifungal supernatant in order to determine the molecular targets involved in inhibtion.
Project description:Microarrays were used to identify genes that participate in the acid responce tolerance of C. jejuni. Three biological replicates of two different conditions (pH 5.5 and 7). Two slides and every slide contained three arrays- see attached additional file.
Project description:Breast Cancer (BC) remains a major cause of mortality, and early detection is considered important for reducing BC-associated deaths. Early detection of BC is challenging in young women, due to the limitations of mammography on the dense breast tissue of young women. We recently reported results of a pilot study, using one-dimensional polyacrylamide gel electrophoresis (1D-PAGE)-based mass spectrometry (MS) to investigate differences in milk proteins from women with and without BC. Here, we applied two-dimensional polyacrylamide gel electrophoresis (2D-PAGE)-based MS to compare the protein pattern in milk from the breasts of a single woman who was diagnosed with BC in one breast 24 months after donating her mil. Statistically different gel spots were picked for protein digestion followed by nanoLC-MS/MS analysis. The upregulated proteins in BC versus control are alpha-amylase, gelsolin isoform a precursor, alpha-2-glycoprotein 1 zinc isoform CRA_b partial, apoptosis-inducing factor 2 and vitronectin. Several proteins were downregulated in the milk of the breast later diagnosed with cancer as compared to the milk from the healthy breast, including different isoforms of albumin, cholesterol esterase, different isoforms of lactoferrin, different proteins from the casein family and different isoforms of lysozyme. Results warrant further studies to determine the usefulness of these milk proteins for assessing risk and detecting occult disease.
Project description:The altitude gradient limits the growth and distribution of alpine plants.Alpine plants have developed strategies to survive the extremely cold conditions prevailing at high altitudes; however, the mechanism underlying the evolution of these strategies remains unknown. The alpine plant Potentilla saundersiana is widespread in the Northwestern Tibetan Plateau. In this study, we conducted a comparative proteomics analysis to investigate the dynamic patterns of protein expression of P. saundersiana located at five different altitudes. We detected and functionally characterized 118 differentially expressed proteins. Our study confirmed that increasing levels of antioxidant proteins, and their respective activities, and accumulation of primary metabolites, such as proline and sugar, confer tolerance to the alpine environment in P. saundersiana. Proteins species associated with the epigenetic regulation of DNA stability and post-translational protein degradation were also involved in this process. Furthermore, our results showed that 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 strictosidine. Based on these findings, we conclude that P. saundersiana uses multiple strategies to adapt to the high-altitude environment of the Northwestern Tibetan Plateau.
Project description:In this study, the cell wall proteomes of 3 different regions of alfalfa stems were compared. These three regions correspond to three phases in the stem development. The sequential extraction of cell wall proteins with CaCl2, EGTA and LiCl-complemented buffers was combined with a gel-based proteome approach and multivariate analysis. Although the highest similarities were observed between the apical and intermediate stem regions, the three proteome patterns are characteristic for each region. In the basal stem region, proteins that bind carbohydrates and have proteolytic activity, as well as enzymes involved in glycan remobilization, accumulate. Beta-amylase and ferritin likewise accumulate more in the basal stem segment. Therefore remobilization of nutrients appears to be an important process in the oldest stem segment. The intermediate and the apical region are sites of cell wall polymer remodelling, as suggested by the high abundance of proteins involved in the remodelling of the cell wall such as xyloglucan endoglucosylase, polygalacturonase non-catalytic subunit or beta-galactosidase. The most striking change between the different stem parts is however the strong accumulation of a DUF642-conserved domain containing protein in the apical region of the stem which suggests a particular role of this protein during the early development of stem tissues.
Project description:Pharmaceuticals are pseudo persistent aquatic pollutants with unknown effects at environmentally relevant concentrations. Atlantic salmon (Salmo salar) were exposed to Acetaminophen: 54.77 ± 34.67; Atenolol: 11.08 ± 7.98 and Carbamazepine: 7.85 ± 0.13 µg•L-1 for 5 days. After Acetaminophen treatment, 19 proteins were differently expressed, of which 11 were significant with respect to the control group (eight up-regulated and three down-regulated). After Atenolol treatment, 7 differently expressed proteins were obtained in comparison with the control, of which 6 could be identified (four up-regulated and 2 down-regulated). Carbamazepine exposure resulted in 15 differently expressed proteins compared with the control, with 10 of them identified (seven up-regulated and three down-regulated). Out of these, 3 features were common between Acetaminophen and Carbamazepine and one between Carbamazepine and Atenolol. One feature was common across all treatments. Principal component analysis and heat map clustering showed a clear grouping of the variability due to the applied treatments. The obtained data suggest (1) that exposure to environmentally relevant concentrations of the pharmaceuticals alters the hepatic protein expression profile of the Atlantic salmon; and (2) the existence of treatment specific processes that may be useful for biomarker development.
Project description:Advances in chemoproteomic technology have revealed covalent interactions between small molecules and protein nucleophiles, primarily cysteine, on a proteome-wide scale. Most chemoproteomic screening approaches are indirect, relying on competition between electrophilic fragments and a minimalist electrophilic probe with inherently limited proteome coverage. Here, we develop a chemoproteomic platform for direct electrophile-site identification based on enantiomeric pairs of clickable arylsulfonyl fluoride probes. Using stereoselective site modification as a proxy for ligandability in intact cells, we identified 634 tyrosines and lysines within functionally diverse protein sites, liganded by structurally diverse probes. Among multiple validated sites, we discovered a chiral probe that modifies Y228 in the MYC binding site of the epigenetic regulator WDR5, as revealed by a high-resolution crystal structure. A distinct chiral probe stimulates tumor cell phagocytosis by covalently modifying Y387 in the recently discovered immuno-oncology target, APMAP. Our work provides a deep resource of ligandable tyrosines and lysines for the development of covalent chemical probes.
Project description:N. benthamiana plants were grown under 16 hour light/8 hour dark cycle in a plant growth room at 24°C for approximately six weeks before subjected to virus-induced gene silencing. Agrobacterium stain GV2260 (OD600=1.0) carrying silencing constructs were infiltrated into 2 fully expended leaves for inducing gene silencing. Samples were collected at 8, 10, 12, 14, and 16 days post Agro-inoculation (DPI) for RPN9-silenced plants and N-silenced control. For RPN8-silencing, samples were collected at 8, 10, and 12 DPI, and the empty vector treated plants were used as a control. Each sample was a pool of 6 silenced leaves collected from 3 individual plants. All the samples were in biological triplicates from 3 sets of independently silenced plants. Total RNA was extracted using Trizol and DNA was removed with DNase I treatment before cDNA synthesis. Keywords: Reference design 25 hybs total
Project description:Data analysis. Spot detection and matching were performed with a comparative cross analysis of all the gels using DeCyder software v.6.5 (GE Healthcare). 178 spots were selected based on 1.15-fold for protein ratio cut-off, allowing for the appearance of the spots in 23 out of 28 gels (69 out of 84 total images). Data from 95 spots were submitted. 93 spots were identified with high confidence. Spot picking and Trypsin digestion. The spots of interest were picked up by Ettan Spot Picker (GE Healthcare) based on the in-gel analysis and spot picking design by DeCyder software. The gel spots were washed a few times then digested in-gel with modified porcine trypsin protease (Promega, Fitchburg, WI). The digested tryptic peptides were desalted using a Zip-tip C18 (Millipore, Billerica, MA). Peptides were eluted from the Zip-tip with 0.5 uL of matrix solution (alpha-cyano-4-hydroxycinnamic acid, 5 mg/mL in 50% acetonitrile, 0.1% trifluoroacetic acid, 25mM ammonium bicarbonate) and spotted on a MALDI plate. Mass Spectrometry. MALDI-TOF MS and TOF/TOF tandem MS/MS were performed on AB SCIEX TOF/TOF 5800 System (AB SCIEX). MALDI-TOF mass spectra were acquired in reflectron positive ion mode, averaging 4000 laser shots per spectrum. TOF/TOF tandem MS fragmentation spectra were acquired for each sample, averaging 4000 laser shots per fragmentation spectrum on each of the 7-10 most abundant ions present in each sample (excluding trypsin autolytic peptides and other known background ions). Database search. Both the resulting peptide mass and the associated fragmentation spectra were submitted to GPS Explorer workstation equipped with MASCOT search engine (Matrix Science, Boston, MA) to search the Swiss-Prot database. Searches were performed without constraining protein molecular weight or isoelectric point, with variable carbamidomethylation of cysteine and oxidation of methionine residues, and with one missed cleavage also allowed in the search parameters. Candidates with either protein score C.I.% or Ion C.I.% greater than 95 were considered significant. When multiple IDs were significant for a given spot, the selection was made by evaluating apparent molecular weight, isoelectric point, the location of the spot in the gel, and the presence of strips of multiple protein isoforms in the adjacent spots.