Project description:Cellular metabolism controls gene expression through allosteric interactions between metabolites and transcription factors. Methods to detect these regulatory interactions are mostly based on in vitro binding assays, but there are no methods to identify them at a genome-scale in vivo. Here we show that dynamic transcriptome and metabolome data identify metabolites that are potential effectors of transcription factors in E. coli. By switching the culture conditions between starvation and growth for 20 hours, we induced strong metabolite concentration changes and accompanying gene expression changes, which were measured by LC-MS/MS and RNA sequencing. From the transcriptome data we calculated the activity of 209 transcriptional regulators with Network Component Analysis, and then tested which metabolites correlated with these activities. This approach captured, for instance, the in vivo Hill-kinetics of CRP regulation by cyclic-AMP, a canonical example of allosteric transcription factor regulation in E. coli. By testing correlations between all pairs of transcription factors and metabolites, we predicted putative effectors of 65 transcription factors, and validated five of them in vitro. These results show that the combination of transcriptomics and metabolomics can generate hypotheses about metabolism-transcription interactions that are relevant in vivo and drive transitions between physiological states.
Project description:The goal was to determine the chemotherapy-induced, mammalian cell death-dependent transcriptional response in a human commensal strain of E. coli. Murine intestinal epithelial cells were used to induce chemotherapy-driven cell death, and a human commensal strain of E. coli was used as the 'recipient' bacteria.
Project description:We studied the response to infection and associated perturbations to the bovine liver’s normal function by examining gene transcription data from liver biopsies collected following an E. coli infection in the udder of primiparous dairy cows. This is the first study to examine gene transcription responses to systemic infection by the E. coli bacterium in dairy cows. First, we verified that the inoculation protocol resulted in systemic infection in the cows. This was done based on records on three clinical symptoms: body temperature and amount of E. coli bacteria and leukocytes in milk samples. Second, we examined gene transcription patterns underlying the clinical traits. Gene transcription levels at times of peak values for the clinical traits were estimated in the liver to study indications of an acute phase response to systemic E. coli infection in the cows. Finally, we compared gene transcription responses to E. coli infection and lipopolysaccaride (LPS) inoculation.
Project description:Brown adipose tissue (BAT) dissipates energy and promotes cardio-metabolic health4. However, loss of BAT during obesity and aging is a principal hurdle for BAT-centered obesity therapies. So far not much is known about BAT apoptosis and signals released by apoptotic brown adipocytes. Here, untargeted metabolomics demonstrated that apoptotic brown adipocytes release a specific pattern of metabolites with purine metabolites being highly enriched. Interestingly, this apoptotic secretome enhances expression of the thermogenic program in healthy adipocytes to maintain tissue functionality. This effect is mediated by the purine inosine which stimulates energy expenditure (EE) in brown adipocytes. Phosphoproteomic analysis demonstrated activation of the cAMP/protein kinase A signaling pathway and of pro-thermogenic transcription factors by inosine.
Project description:Apoptosis accounts for ~ 90% of homeostatic cell turnover in the body1. While caspase-dependent apoptosis is known to influence immune tolerance, induction of cell proliferation, and tissue regeneration2-4, how these apoptotic cells contribute to such diverse effects is less understood. Soluble factors released from apoptotic cells could be a means of ‘talking’ to healthy cells in the tissue neighborhood. While a few phagocyte-attracting ‘find-me’ signals released from apoptotic cells are known5, the larger ‘metabolite secretome’ from apoptotic cells is not yet defined. Here, via unbiased profiling of the apoptotic cell secretome, we identified 123 metabolites that are specifically released during apoptosis, while cells still retain membrane integrity. Release of ~25 of these metabolites is dependent on Pannexin 1 (Panx1) channels, which are opened by caspase-dependent cleavage during apoptosis. Interestingly, one of the Panx1-dependent metabolites from apoptotic cells is spermidine, a polyamine whose release could be, in part, due to the continued metabolic activity of dying cells. RNAseq analysis of healthy cells exposed to the apoptotic secretome revealed Panx1-dependent gene programs, including genes linked to suppression of inflammation, cell proliferation, and wound healing. Analysis of the apoptotic secretome across cell types and different apoptotic stimuli, identified a core “set” of 8 Panx1-dependent metabolites. Strikingly, a selected cocktail of these metabolites demonstrated significant immunosuppressive effects by potently reducing disease severity in an inflammatory arthritis model and a lung graft rejection model. Collectively, these data advance the concept that apoptotic cells are not ‘inert’ corpses waiting for removal by phagocytes, but rather they actively communicate with the tissue environment through their metabolite secretome, with implications in tissue inflammation.
Project description:Environmental fluctuations lead to a rapid adjustment of the physiology of Escherichia coli, necessitating changes on every level of the underlying cellular and molecular network. Thus far, the vast majority of global analyses of E. coli stress responses have been limited to just one level, gene expression. Here we incorporate the metabolite composition together with gene expression data in order to provide a more comprehensive insight on system level stress adjustments by describing detailed time-resolved E. coli response to five different perturbations (cold, heat, oxidative stress, lactose diauxie, and stationary phase). The metabolite response is more specific as compared to the general response observed on the transcript level and is reflected by much higher specificity during the early stress adaptation phase and when comparing the stationary phase response to other perturbations. Despite these differences, the response on both levels still follows the same dynamics and general strategy of energy conservation as reflected by rapid decrease of central carbon metabolism intermediates coinciding with down regulation of genes related to cell growth. Application of co-clustering and canonical correlation analysis on combined metabolite and transcript data identified a number of significant condition dependent associations between metabolites and transcripts. The results confirm and extend existing models about co-regulation between gene expression and metabolites demonstrating the power of integrated systems oriented analysis.
Project description:E. coli O157:H7 can use vegetables such as spinach as secondary hosts, from where they can be transmitted into the food chain. Our previous microarray analysis showed whole-scale changes in gene expression of E. coli driven in a large part in response to plant metabolites. The aim of this work was to expand beyond the inherent limitations of microarray analysis, taking an RNA-seq approach to analyse the same samples.