Valine-Induced Isoleucine Starvation in Escherichia coli K-12 Studied by Spike-In Normalized RNA Sequencing
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ABSTRACT:
PROVIDER: PRJNA563584 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR10059464_1.fastq.gz | Fastqsanger.gz | |||
| SRR10059464_2.fastq.gz | Fastqsanger.gz | |||
| SRR10059465_1.fastq.gz | Fastqsanger.gz | |||
| SRR10059465_2.fastq.gz | Fastqsanger.gz | |||
| SRR10059466_1.fastq.gz | Fastqsanger.gz |
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Project description:Here we report on an RNAseq method in combination with spike-in cells to measure global changes in the transcription pattern after valine-induced isoleucine starvation of a standard E. coli K12 strain. Due to the spike-in method we were able to show that ribosomal RNA is degraded during isoleucine starvation and we showed how this change in cellular RNA content affect the estimated regulation of mRNA levels compared to if spike-in were not utilized. Our analysis also showed that induction of starvation by sudden addition of high valine concentrations provoked prominent regulatory responses outside of the expected ppGpp, RpoS and Lrp regulons
2019-11-19 | GSE136753 | GEO
Project description:Escherichia coli cells respond to a period of famine by globally reorganizing their gene expression. The changes are known as the stringent response, which is orchestrated by the alarmone ppGpp that binds directly to RNA polymerase. The resulting changes in gene expression are particularly well studied in the case of amino acid starvation. We used deep RNA sequencing in combination with spike-in cells to measure global changes in the transcriptome after valine-induced isoleucine starvation of a standard E. coli K12 strain. Owing to the whole-cell spike-in method that eliminates variations in RNA extraction efficiency between samples, we show that ribosomal RNA levels are reduced during isoleucine starvation and we quantify how the change in cellular RNA content affects estimates of gene regulation. Specifically, we show that standard data normalization relying on sample sequencing depth underestimates the number of down-regulated genes in the stringent response and overestimates the number of up-regulated genes by approximately 40%. The whole-cell spike-in method also made it possible to quantify how rapidly the pool of total messenger RNA (mRNA) decreases upon amino acid starvation. A principal component analysis showed that the first two components together described 69% of the variability of the data, underlining that large and highly coordinated regulons are at play in the stringent response. The induction of starvation by sudden addition of high valine concentrations provoked prominent regulatory responses outside of the expected ppGpp, RpoS, and Lrp regulons. This underlines the notion that with the high resolution possible in deep RNA sequencing analysis, any different starvation method (e.g., nitrogen-deprivation, removal of an amino acid from an auxotroph strain, or valine addition to E. coli K12 strains) will produce measurable variations in the stress response produced by the cells to cope with the specific treatment.
| S-EPMC7066862 | biostudies-literature
Project description:Abstract: Non-alcoholic steatohepatitis (NASH) is associated with a disturbed metabolism in liver and excessive accumulation of ectopic fat. Branched-chain amino acids (BCAAs) may beneficially modulate hepatic lipids, however it remains unclear whether individual BCAAs can attenuate already established NASH and oxidative-inflammatory stress associated with it. Therefore, mice were first put on a run-in fast food diet (FFD) for 26 weeks, these obese mice were then treated with individual BCAAs valine or isoleucine (3% of FFD) for another 12 weeks and compared to FFD controls. Valine and isoleucine treatment did not affect obesity, dyslipidemia, gut permeability or fecal fatty acid excretion, although significantly reduced hyperinsulinemia compared with FFD. Valine and isoleucine significantly reduced ALT, CK-18M30, and liver steatosis with a particularly pronounced effect on the microvesicular component (-61% by valine and -71% by isoleucine). Hepatic 4-hydroxynonenal (4-HNE) immunoreactivity, a marker for oxidative stress-induced lipid peroxidation, was also significantly decreased. Functional genomics analysis demonstrated that valine and isoleucine upregulated BCAA metabolism, deactivated master regulators of anabolic pathways related to steatosis (e.g. SREBPF1) and activated master regulators of mitochondrial biogenesis (e.g. PPARGC1a) and lipid catabolism (e.g. ACOX1, AMPK). The correction of critical metabolic pathways by valine and isoleucine was accompanied by a significant decrease of liver inflammation, and suppression of many FFD-stimulated cytokine and chemokine pathways including IL-1β, TNFα and CCR2 signaling. In conclusion, the dietary intervention with either valine or isoleucine corrected multiple metabolic processes in liver and reduced steatosis and inflammation with profound effects on oxidative stress and inflammatory pathways. Methods: Putative antifibrotic effects of butyrate were studied in Ldlr-/-.Leiden mice fed an obesogenic NASH-inducing diet (HFD) containing 2.5% (w/w) butyrate for 38 weeks and compared to an HFD control group. Antifibrotic mechanisms of butyrate were further investigated in TGF-β-stimulated primary human hepatic stellate cells (HSC). Results: HFD-fed mice developed obesity, insulin resistance, increased levels of plasma leptin, adipose tissue inflammation, gut permeability, dysbiosis and NASH-associated fibrosis. Butyrate corrected hyperinsulinemia, lowered plasma leptin levels and attenuated adipose tissue inflammation, without affecting gut permeability or microbiota composition. Butyrate lowered plasma ALT and CK-18M30 and attenuated hepatic steatosis and inflammation. Butyrate inhibited fibrosis development as demonstrated by decreased hepatic collagen content and Sirius-red-positive area. In TGF-β-stimulated HSC, butyrate dose-dependently reduced collagen deposition and decreased procollagen1α1 and PAI-1 protein expression. Transcriptomic analysis and subsequent pathway and upstream regulator analysis revealed deactivation of specific non-canonical TGF-β signaling pathways RhoA/Rock and PI3K/AKT and other important pro-fibrotic regulators (e.g. YAP/TAZ and MYC) by butyrate, providing a potential rationale for its antifibrotic effects. Conclusions: Butyrate protects against the development of obesity and insulin resistance-associated NASH and liver fibrosis. These antifibrotic effects are at least in part attributable to a direct effect on collagen production in hepatic stellate cells, as a result of inhibiting non-canonical TGF-β signaling.
2022-06-30 | GSE195798 | GEO
Project description:For the first time in Lactococcus lactis, amino acid starvation response was characterized. The natural imposition of isoleucine starvation, by its consumption during growth, associated to transcript profiling, allowed defining exhaustively this stress stimulon. It consisted of a general induction of nitrogen metabolism (amino acid biosynthesis and transport, proteolytic system and proteases), a strong repression of genes encoding major physiological activities (translation, transcription, carbon metabolism, purine and pyrimidine biosynthesis and fatty acid metabolism) and the induction of unexpected cross responses to acid, osmotic and oxidative stresses. Keywords: stress response, time course Isoleucine starvation was imposed by the consumption of this amino acid during the growth of Lactococcus lactis IL1403 on ILV0.1 medium (CDM with ten-fold reduced concentrations of isoleucine, leucine and valine) and under controlled conditions (30 °C, pH 6.6, nitrogen atmosphere). Cell samples were harvested in exponential phase and after 30 min, 1.7 h and 3.5 h of isoleucine starvation. Total RNA was extracted from these samples and radiolabelled cDNA were prepared and hybridized on nylon arrays. 2053 amplicons specific of Lactococcus lactis IL1403 genes were spotted twice on the array. The 4 time-points were analyzed simultaneously and 3 independent repetitions were performed.
2010-06-25 | E-GEOD-4872 | biostudies-arrayexpress
Project description:ChIP-seq for Leu3 with addition of different levels of leucine, valine and isoleucine
| PRJEB36934 | ENA
Project description:Colorectal cancer (CRC) is currently the third in cancer incidence worldwide and the fourth most common cause of cancer deaths. To discover the proteins related to colon cancer, a typical inflammation-related C57B/6N mouse colon carcinogenesis model was developed using azoxymethane-dextran sodium sulfate (AOM-DSS) treated for 14 weeks. iTRAQ-based proteomics study was performed using cell membrane components enriched from colonic mucosa. Tumor tissues and their adjacent normal colon tissues from colonic cancer patients were collected for differential protein detection and metabolomics studies. Totally, 74 differentially expressed proteins were identified in the AOM/DSS treated tumor samples compared with AOM/DSS treated adjacent samples, and the saline treated control. Bioinformatics analysis showed that eight downregulated proteins were enriched in the pathway of valine, leucine and isoleucine degradation. Targeted metabolomics study showed that valine, leucine and isoleucine was upregulated in tumor tissues compared with their adjacent normal tissues from colonic cancer patients. Further real-time PCR and western blot experiments showed that the signal pathway proteins of Hadh and ALDH2 were downregulated in colon patients (colon tumor tissues vs their adjacent colon tissues), as well as in AOM/DSS treated mouse model. In all, our study showed that the pathway of valine, leucine and isoleucine degradation was inactivation in colon cancer, and Hadh and ALDH2 were two potential biomarkers for colon cancer treatment.
2023-11-13 | PXD046882 |
Project description:Isoleucine and valine correct hepatic lipid processing, reduce liver steatosis and suppress inflammation in obese mice with manifest NASH
| PRJNA802192 | ENA
Project description:The purpose was to find changes in the E. coli transcriptome as a result of isoleucine starvation. rRNA depletion at the library preparation stage was avoided so changes in levels of any ncRNAs could be gauged as accurately as possible. Samples were sequenced at added depth to make up for lack of rRNA depletion.
2017-03-31 | E-MTAB-5221 | biostudies-arrayexpress
Project description:Amino acid assimilation and metabolism are crucial for bacterial growth and survival and this is particularly obvious for lactic acid bacteria (LAB) that are generally auxotroph for various amino acids. However, amino acid assimilation is poorly characterized and a complete description of the response during amino acid starvation is still lacking in LAB. In this context, the global response of the LAB model Lactococcus lactis was characterized during isoleucine starvation in batch culture. The stress was imposed by isoleucine natural consumption in an initially rich chemically defined medium. Dynamic analyses were performed both using transcriptomic and proteomic approaches. The response was found to occur gradually and could be divided into three major parts that were firstly deduced from transcriptomic analysis and generally corroborated by proteomic results: (i) a global repression of biogenic processes (transcription, translation, and carbon metabolism and transport), (ii) a specific response related to the limiting nutrient (numerous pathways belonging to carbon or nitrogen metabolism and leading to isoleucine supply were activated) and (iii) an additional response connected to oxidative stress (induction of aerobic metabolism, electron transport, thioredoxin metabolism and pyruvate dehydrogenase). The involvement of various regulatory mechanisms such as growth rate regulation, stringent response, CodY, GlnR, and CcpA regulations, was discussed on the basis of transcriptomic data comparisons. Above the full description of L. lactis isoleucine starvation response, this work additionally provided a complex but realistic outlook of the regulation network involved in isoleucine starvation. Such integrated and comparative approach will allow, by its implementation to other regulations and environmental conditions, the whole regulatory network of L. lactis or any other microorganism to be deciphered.
2011-09-22 | GSE12962 | GEO
Project description:To characterize the differentially expressed genes between adding Valine and without Valine on Streptomyces natalensis HW-2
2018-07-15 | GSE116624 | GEO