Project description:We studied the proteome changes in Escherichia coli growing in rich media, and compared the changes between exponential phase cells and those underwent sudden carbon starvations.
Project description:Protein lysine methylation is a common post-translational modification (PTM) detected throughout the human proteome that plays important roles in diverse biological processes. In the human genome, there are greater than one hundred known and candidate protein lysine methyltransferases (PKMTs), many with important links to human disease. METTL21C (methyltransferase-like protein 21C) is a PKMT implicated in muscle biology and reported to methylate VCP (valosin-containing protein/p97) and HSPA8 (heat shock 70kDa protein 8). However, a clear in vitro methyltransferase activity for METTL21C is yet to be demonstrated. Thus, whether METTL21C is indeed an active enzyme that directly methylates substrate/s is unclear. Here, we use an unbiased biochemical-based screening assay coupled to mass spectrometry to identify Alanine-tRNA-Synthetase 1 (AARS1) as a direct substrate of METTL21C. METTL21C catalyzes mono-, di- and tri-methylation of lysine 943 of AARS1 (AARS1-K943me) in vitro and in vivo. In vitro methylation of AARS1 by METTL21C is independent of ATP or tRNA molecules. In contrast to AARS1, and in conflict with previous reports, we do not detect METTL21C methylation activity on VCP and HSPA8. Depletion of METTL21C in METTL21C-expressing cells leads to depletion of AARS1-K943 methylation. Finally, METTL2C is almost exclusively expressed in muscle tissue, and accordingly we detect the METTL21C-catalyzed methylation event of AARS1 in mouse skeletal muscle tissue. Together, our work identifies AARS1 as a bona fide substrate of METTL21C and suggests a role for the METTL21C-AARS1 axis in the regulation of protein synthesis in muscle tissue. In addition, our study describes a straightforward protocol for elucidating physiologic substrates of little characterized or uncharacterized PKMTs
Project description:To study the effect of microbe-associated molecular pattern (MAMP) treatment (flg22) on the phosphorylation of nuclear proteins, we treated Arabidopsis plants with flg22 and after isolation of nuclear proteins, we enriched for phosphopeptides and then carried out LC-MS/MS analyses. We used statistical analysis tools to identify differentially phosphorylated proteins in response to MAMP treatment in the mock and treated samples.
Project description:Assessment of individual whole cell transcriptional variation among closely related bacterial ecotypes Here we used 4 replicates each containing 3 independent biological replicates as well as dye swaps per each replicate. We hybridized in a two color format but analyzed in a one color format.
Project description:Glycolysis is the central pathway for sugar metabolism in most living organisms. The single celled eukaryote yeast is a widely used model organism for higher eukaryotes in which the regulation of glycolysis is widely studied. S. cerevisiae is one of the few eukaryotic organisms that can efficiently grow under both aerobic and anaerobic conditions. Furthermore, this yeast switches from proliferation into a resting phase when nutrients are exhausted. However, little is known about the proteome dynamics that takes place during this transition, particularly under anaerobic conditions. Moreover, like many other organisms including humans, the genome of S. cerevisiae contains duplications which result in the expression of so called ‘isoenzymes’ in central metabolic pathways including glycolysis. Interestingly, the role of those ‘isoenzymes’ remains elusive to date. Here we describe a large-scale quantitative proteome study combining shotgun experiments using a nano-LC coupled to a QE plus Orbitrap mass spectrometer (Thermo, Germany), to capture the proteome dynamics during transition from proliferation into stationary phase, under both aerobic and anaerobic growth. Furthermore, we explore the proteome dynamics of a mutant yeast where the glycolytic isoenzymes are deleted.
Project description:Naturally bacteria are commonly forced to remain in stationary phase. There is no increase in cell mass, however, cell division keep on. Vibrio (V.) parahaemolyticus is an aquatic bacterium capable of causing foodborne gastroenteritis outbreaks all over the world. So far, little is known about whole genomic expression of V. parahaemolyticus in the early stationary phase compared with the phase of exponential growth. Since under starvation cell sizes decrease and endogenous metabolism reduces, genes are considered to be highly repressed in the stationary phase. However, our data shows in total 172 induced genes, while 61 genes were repressed in the early stationary phase compared with exponential phase. In fatty acid and phospholipid metabolism functional category only induced genes were found, whereas in three other metabolic functional groups appeared no significant up-regulated genes (adjusted P-value<0.05). Genes in two metabolic functional categories remained stable in the early stationary phase. DAVID analyses were carried out exploring the gene regulation. In total, ten functional categories showed a total up-regulation in early stationary phase, while only three metabolic functional categories showed a down-regulation and four categories showed stably in early stationary phase. Early stationary phase gene expression was detected in total bacterial RNA of V. parahaemolyticus. Two phases (exponential phase and early stationary phase) were used in 8 biological replicates. Gene expression in exponential phase was used for normalization.
Project description:To investigate the influence of chromatin organization and dynamics on the response to Notch signaling, we partitioned Drosophila chromatin using histone modifications and established This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
Project description:SARS-CoV-2 nsp7 and nsp8 are important cofactors of the RTC, as they interact and regulate the activity of RNA-dependent RNA polymerase and other nspsHere we used solution-based structural proteomic techniques, hydrogen-deuterium exchange mass spectrometry (HDX-MS) and crosslinking mass spectrometry (XL-MS), illuminate the dynamics of SARS-CoV-2 full-length nsp7, nsp8, and nsp7:nsp8 proteins and protein complex.
Project description:Circular RNAs (circRNA) are a novel class of widespread non-coding RNAs (ncRNAs) that regulate gene expression in mammals. Recent studies demonstrate that functional peptides can be encoded by short open reading frames (sORFs) in ncRNAs, including circRNAs. In this study, through deep RNA sequencing on human endometrial cancer (EC) samples and their paired adjacent normal tissues, we uncovered that the circRNA hsa-circ-0000437 is significantly reduced in EC compared to matched paracancerous tissue. The hsa-circ-0000437 contains a sORF encoding a functional peptide termed as CORO1C-47aa. Overexpression of CORO1C-47aa is capable of inhibiting angiogenesis at the initiation stage by suppressing endothelial cell proliferation, migration, and differentiation through competing with TACC3 to bind to ARNT and suppress VEGF. The anti-tumor effects of CORO1C-47aa on EC progression suggest that CORO1C-47aa has a potential value in anti-carcinoma therapies and deserves further investigation.
Project description:The MalNO is a putative two-component signal transduction system, previously known as the VirJI sytem. MalO is the putative cognate response regulator of the MalN sensor histidine kinase. Based on previous evidence that suggested the plc gene, encoding ?-toxin, was upregulated during stationary phase in the malO mutant, microarrays were used to analyse the transcriptome of a malO mutant during stationary phase growth. Total RNA was isolated from stationary phase cells of the malO mutant and the wild-type control. Gene expression levels were compared between the malO mutant and wild-type strain 13