Project description:The cell cycle is predominantly controlled by Cyclins/Cyclin-Dependent Kinases (Cyc/CDK) complexes, which phosphorylate targets involved in cellular proliferation. Evidence suggests that Cyc/CDK targets extend beyond traditional proteins and include enzymes that regulate the central carbon metabolism. Maize embryo axes rapidly internalize and metabolize glucose. After 24 h of imbibition in glucose-rich media, axes exhibited increased length and weight, with more pronounced effects at 72 h. This morphology enhancement was impaired when RO-3306, a specific CDK inhibitor, was added. The protein profile of maize embryo extracts at 18 and 24 h indicated altered phosphorylation patterns following CDK activity inhibition. Metabolomic analysis at 24 h of imbibition revealed that maize embryos without sugar in the media, with or without RO-3306, had a decreased sugar and amino acid content. Conversely, axes exposed to glucose demonstrated increased conversion into various mono and di-saccharides such as fructose, mannitol, galactose, and maltose but not sucrose. This pattern was reversed upon the addition of RO-3306. Glucose promoted the accumulation of amino acids such as cysteine, valine, leucine, and intermediates of the tricarboxylic acid (TCA) cycle, such as malate and citrate. The CDK inhibitor redirected the glucose metabolism toward increased serine levels, followed by other amino acids like phenylalanine, valine, and leucine. Additionally, TCA cycle intermediates and sterols significantly decreased. Overall, these results contribute to understanding the role of CDK in maize morphogenesis during germination and underscore its impact on modulating various central carbon pathways, including glycolysis, amino acid catabolism/anabolism, TCA cycle, and sterols biosynthesis.
Project description:The fast growing evidences have shown that the plant-derived compound honokiol is a promising candidate for treating multiple human diseases, such as inflammation and cancer. However, the mode-of-action (MoA) of honokiol remains largely unclear. Here, we studied the antifungal activity of honokiol in fission yeast model, with the goal of understanding the honokiol's mechanism of action from the molecular level. We found that honokiol can inhibit the yeast growth at a dose-dependent way. Microarray analysis showed that honokiol has wide impacts on the fission yeast transcription levels (in total, 512 genes are up-regulated, and 42 genes are down-regulated). Gene set enrichment analysis indicated that over 45% up-regulated genes belong to the core environmental stress responses category. Moreover, network analysis suggested that there are extensive gene-gene interactions amongst the co-expression gene lists, which can assemble several biofunctionally important modules. It is noteworthy that several key components of central carbon metabolism, such as glucose transporters and metabolic enzymes of glycolysis, are involved in honokiol's MoA. The complexity of the honokiol's MoA displayed in previous studies and this work demonstrates that multiple omics approaches and bioinformatics tools should be applied together to achieve the complete scenario of honokiol's antifungal function.
Project description:We used Progenika oligonucleotide arrays to monitore the gene expression of P. putida during carbon source stimulus experiments. After ensuring steady state conditions on benzoate, the stimuli were introduced by changing the carbon source from benzoate to glucose, from glucose to fructose and from fructose to benzoate, respectively. The single stimuli were monitored over a time period from 10 to 120 minutes after changing the carbon source by three representative timepoints. Moreover, the steady state conditions were compared among each other. Supplementary file: Individual normalized signal intensity VALUES for each channel of each array are provided in the FullNormalizedMatrix.txt file.
Project description:As a result of ancestral whole genome and small-scale duplication events, the genome of Saccharomyces cerevisiaeM-bM-^@M-^Ys, and of many eukaryotes, still contain a substantial fraction of duplicated genes. In all investigated organisms, metabolic pathways, and more particularly glycolysis, are specifically enriched for functionally redundant paralogs. In ancestors of the Saccharomyces lineage, the duplication of glycolytic genes is purported to have played an important role leading to S. cerevisiae current lifestyle favoring fermentative metabolism even in the presence of oxygen and characterized by a high glycolytic capacity. In modern S. cerevisiae, the 12 glycolytic reactions leading to the biochemical conversion from glucose to ethanol are encoded by 27 paralogs. In order to experimentally explore the physiological role of this genetic redundancy, a yeast strain with a minimal set of 14 paralogs was constructed (MG strain). Remarkably, a combination of quantitative, systems approach and of semi-quantitative analysis in a wide array of growth environments revealed the absence of phenotypic response to the cumulative deletion of 13 glycolytic paralogs. This observation indicates that duplication of glycolytic genes is not a prerequisite for achieving the high glycolytic fluxes and fermentative capacities that are characteristic for S. cerevisiae and essential for many of its industrial applications and argues against gene dosage effects as a means for fixing minor glycolytic paralogs in the yeast genome. MG was carefully designed and constructed to provide a robust, prototrophic platform for quantitative studies, and is made available to the scientific community. The goals of the present study are to experimentally explore genetic redundancy in yeast glycolysis by cumulative deletion of minor paralogs and to provide a new experimental platform for fundamental yeast research by constructing a yeast strain with a functional M-bM-^@M-^Xminimal glycolysisM-bM-^@M-^Y. To this end, we deleted 13 minor paralogs, leaving only the 14 major paralogs for the S. cerevisiae glycolytic pathway. The cumulative impact of deleting all minor paralogs was investigated by two complementary approaches. A first, quantitative analysis focused on the impact on glycolytic flux under a number of controlled cultivation conditions that, in wild-type strains, result in different glycolytic fluxes. These quantitative growth studies were combined with transcriptome, enzyme-activity and intracellular metabolite assays to capture potential small phenotypic effects. A second, semi-quantitative characterization explored the phenotype of the M-bM-^@M-^Xminimal glycolysisM-bM-^@M-^Y strain under a wide array of experimental conditions to identify potential context-dependent phenotypes
Project description:Cyclic diadenosine monophosphate (c-di-AMP) is a conserved nucleotide second messenger critical for bacterial growth and resistance to cell wall-active antibiotics. In Listeria monocytogenes, the sole diadenylate cyclase, DacA, is essential in rich, but not synthetic media and ΔdacA mutants are highly sensitive to the β-lactam antibiotic cefuroxime. In this study, loss of function mutations in the oligopeptide importer (oppABCDF) and glycine betaine importer (gbuABC) allowed ΔdacA mutants to grow in rich medium. Since oligopeptides were sufficient to inhibit growth of the ΔdacA mutant we hypothesized that oligopeptides act as osmolytes, similar to glycine betaine, to disrupt intracellular osmotic pressure. Supplementation with salt stabilized the ΔdacA mutant in rich medium and restored cefuroxime resistance. Additional suppressor mutations in the acetyl-CoA binding site of pyruvate carboxylase (PycA) rescued cefuroxime resistance and resulted in a 100-fold increase in virulence of the ΔdacA mutant. PycA is inhibited by c-di-AMP and these mutations prompted us to examine the role of TCA cycle enzymes. Inactivation of citrate synthase, but not down-stream enzymes suppressed ΔdacA phenotypes. These data suggested that c-di-AMP modulates central metabolism at the pyruvate node to moderate citrate production and indeed, the ΔdacA mutant accumulated six times the concentration of citrate present in wild-type bacteria.
Project description:The field of epitranscriptomics is growing in importance, with chemical modification of RNA being associated with a wide variety of biological phenomena. Mass spectrometry (MS) enables the identification of modified RNA residues within their sequence contexts, by using analogous approaches to shotgun proteomics. We have developed a free and open-source database search engine for RNA MS data, called NucleicAcidSearchEngine (NASE), as part of the OpenMS software framework. NASE allows the reliable identification of (modified) RNA sequences from LC-MS/MS data in a high-throughput fashion. For this validation dataset, oligonucleotides with the sequence of mature Drosophila let-7 microRNA, 21 nt in length, were produced synthetically in unmodified and modified (2’-O-methylated at the 3’ uridine) forms. We characterised a 1:1 mixture of both forms of this RNA.
Project description:Central fatigue is defined as a failure of the central nervous system to adequately drive the muscle, manifesting limited development, and maintenance of locomotor activity. A plateau in hypoxia leads to central fatigue and followed by maximal motility recession. However, the underlying mechanism is still unclear. The present study describes a mechanism by which liver CEBPβ (CCAAT/enhancer-binding protein beta) induced by hypoxic environment alters the kynurenine (KYN) metabolism and causes the suppression of motility function recession. The activation of CEBPβ under hypoxia increases the liver expression of tryptophan dioxygenase, thereby enhancing the conversion of tryptophan into KYN; the KYN metabolite can traverse the blood-brain barrier and result in the suppression of motility function. However, the knockdown of CEBPβ by injecting pAAV-shRNA-CEBPβ via the hepatic portal vein reduces the KYN production and improves the motility function. KYN is a neurochemical that which restricts the exercise capacity after injection in the basal ganglia in mice. Reducing the plasma KYN protects the brain from hypoxia-induced changes associated with fatigue, and the knockdown liver of CEBPβ in mice renders resistance to fatigue post-acute hypoxia or tryptophan treatment. This study reveals resistance to central fatigue as a strategy for acclimatization to hypoxia mediated by transcription factor CEBPβ in the liver.
Project description:The field of epitranscriptomics is growing in importance, with chemical modification of RNA being associated with a wide variety of biological phenomena. Mass spectrometry (MS) enables the identification of modified RNA residues within their sequence contexts, by using analogous approaches to shotgun proteomics. We have developed a free and open-source database search engine for RNA MS data, called NucleicAcidSearchEngine (NASE), as part of the OpenMS software framework. NASE allows the reliable identification of (modified) RNA sequences from LC-MS/MS data in a high-throughput fashion. For this validation dataset, we generated samples of human long ribosomal RNA from a cellular extract. The samples were RNase-treated prior to nanoflow LC-MS/MS analysis.
Project description:The field of epitranscriptomics is growing in importance, with chemical modification of RNA being associated with a wide variety of biological phenomena. Mass spectrometry (MS) enables the identification of modified RNA residues within their sequence contexts, by using analogous approaches to shotgun proteomics. We have developed a free and open-source database search engine for RNA MS data, called NucleicAcidSearchEngine (NASE), as part of the OpenMS software framework. NASE allows the reliable identification of (modified) RNA sequences from LC-MS/MS data in a high-throughput fashion. For this validation dataset, we generated a sample of human total tRNA from a cellular extract - a complex mixture of highly modified RNAs. This sample was RNase-treated prior to nanoflow LC-MS/MS analysis.