Project description:Flavin-binding fluorescent proteins (FPs) are genetically encoded in vivo reporters, which are derived from microbial and plant LOV photoreceptors. In this study, we comparatively analyzed the ROS-induced stress responses of the two variants DsFbFP M49I and Pp2FbFP, exhibiting preferential photosensitization of superoxide and singlet oxygen, respectively, in E.coli.
Project description:In order to study the role of Farnesyl diphosphate synthase (FPS) in post-embryonic plant development, we generated Arabidopsis conditional knockdown mutants expressing amiRNAs devised to simultaneously silence both FPS (FPS1 and FPS2) genes. The expression of the amiRNAs is regulated by using a methoxyfenozide (MFZ) inducible promoter. An RNAseq analysis was made to compare the differentially expressed genes between control (not induced) and silenced (MFZ-induced) amiFPSa Arabidopsis seedlings. Differential gene expression analysis shows that a reduction in FPS activity levels triggers misregulation of genes involved in biotic and abiotic stress responses, the most prominent being the rapid induction of a set of genes related to the jasmonic acid (JA) pathway. Down-regulation of FPS also triggered a Fe-deficiency transcriptional response that is consistent with the Fe-deficient status observed in FPS silenced plants.
Project description:Escherichia coli (E. coli) mazEF is an extensively studied stress-induced toxin-antitoxin (TA) system. The toxin MazF is an endoribonuclease that cleaves RNAs at ACA sites. Thereby, under stress the induced MazF generates a Stress induced Translation Machinery (STM), composed of MazF processed mRNAs and selective ribosomes that specifically translate the processed mRNAs. Here we performed a proteomic analysis of all the E. coli stress-induced proteins that are mediated through chromosomally borne mazF gene . We show that the mRNAs of almost all of them is characterized by the presence of an ACA site up to 100 nucleotides upstream to the AUG initiator. Thereby, under stressful conditions the induced MazF is processing mRNAs that are translated by STM. Furthermore, the presence of the ACA site far enough upstream (up to 100 nucleotides) to the AUG initiator may still permit translation by the canonical translation machinery. Thus, such dual translation mechanisms, enable under stress also to prepare proteins for immediate functions while coming back to normal growth conditions.
Project description:Since the molecular mechanisms behind adaptation and the bacterial stress response toward antimicrobial photodynamic therapy (aPDT) are not entirely clear yet, the aim of the present study was to investigate the transcriptomic stress response in Escherichia coli after sublethal treatment with aPDT using RNA sequencing (RNA-Seq). Planktonic cultures of stationary phase E. coli were treated with aPDT using a sublethal dose of the photosensitizer SAPYR. After treatment, RNA was extracted, and RNA-Seq was performed on the Illumina NextSeq 500. Differentially expressed genes were analyzed and validated by qRT-PCR. The analysis of the differential gene expression following pathway enrichment analysis revealed a considerable number of genes and pathways significantly up- or down-regulated in E. coli after sublethal treatment with aPDT. Expression of 1018 genes was up-regulated and of 648 genes was down-regulated after sublethal treatment with aPDT as compared to irradiated controls. Analysis of differentially expressed genes and significantly de-regulated pathways showed regulation of genes involved in oxidative stress response and bacterial membrane damage. In conclusion, the results show a transcriptomic stress response in E. coli upon exposure to aPDT using SAPYR and give an insight into potential molecular mechanisms that may result in development of adaptation
Project description:Background: The food-borne pathogen Campylobacter is one of the most important zoonotic pathogens. Compared to other zoonotic bacteria, Campylobacter species are quite susceptible to environmental or technological stressors. This might be due to the lack of many stress response mechanisms described in other bacteria. Nevertheless, Campylobacter is able to survive in the environment and food products. Although some aspects of the heat stress response in Campylobacter (C.) jejuni are already known, information about the heat stress response in the related species C. coli and C. lari are still unknown. Results: The stress response to elevated temperatures (46°C) was investigated by survival assays and whole transcriptome analyses for the strain C. jejuni NCTC11168, C. coli RM2228 and C. lari RM2100. While C. jejuni showed highest thermotolerance followed by C. lari and C. coli, none of the strains survived at this temperature for more than 24 hours. Transcriptomic analyses revealed that only 3 % of the genes in C. jejuni and approx. 20 % of the genes of C. coli and C. lari were differentially expressed after heat stress, respectively. The transcriptomic profiles showed enhanced gene expression of several chaperones like dnaK, groES, groEL and clpB in all strains, but differences in the gene expression of transcriptional regulators like hspR, perR as well as for genes involved in metabolic pathways, translation processes and membrane components. However, the function of many of the differentially expressed gene is unknown so far. Conclusion: We could demonstrate differences in the ability to survive at elevated temperatures for C. jejuni, C. coli and C. lari and showed for the first time transcriptomic analyses of the heat stress response of C. coli and C. lari. Our data suggest that the heat stress response of C. coli and C. lari are more similar to each other compared to C. jejuni, even though on genetic level a higher homology exists between C. jejuni and C. coli. This indicates that stress response mechanisms described for C. jejuni might be unique for this species and not necessarily transferable to other Campylobacter species.
Project description:The stress effect of 100 uM sodium tetrachloropalladate (II) was evaluated in Escherichia coli K-12 BW25113 strain on the transcriptome level
Project description:We show that NtrC couples the Ntr stress response and stringent response in N starved E. coli, which appears to be a conserved adaptive strategy employed by many bacteria to manage conditions of nutritional adversity. N starved Escherichia coli initiate the nitrogen regulation (Ntr) stress response as an adaptive mechanism to scavenge for alternative N sources. The Ntr stress response requires the global transcriptional regulator nitrogen regulatory protein C (NtrC). We discovered that the transcription of relA, the key gene responsible for the synthesis of the major effector nucleotide alamorne of the bacterial stringent response, guanosine pentaphosphate (ppGpp), is positively regulated by NtrC in N starved E. coli. we addressed Ntr stress response-ppGpp alarmone links and mapped the genome-wide binding targets of NtrC in E. coli during N starvation using chromatin immunoprecipitation (ChIP) followed by high-throughput sequencing (ChIP-seq) to gain insight into the NtrC-dependent gene networks. To identify candidate genome regions that are preferentially associated with NtrC, we introduced an in-frame fusion encoding three repeats of the FLAG epitope to the 3-prime end of glnG in E. coli strain NCM3722, a prototrophic E. coli K-12 strain.
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:Assimilation of nitrogen is an essential process in bacteria. The nitrogen regulation stress response is an adaptive mechanism used by nitrogen-starved Escherichia coli to scavenge for alternative nitrogen sources and requires the global transcriptional regulator NtrC.
Project description:The response to acidity is crucial for neutralophilic bacteria. Escherichia coli has a well characterized regulatory network to induce multiple defense mechanisms against excess of protons. Nevertheless, systemic studies of the transcriptional and translational reprogramming of E. coli to different acidic strengths have not yet been performed. Here, we used ribosome profiling and mRNA sequencing to determine the response of E. coli to pH 7.6, 5.8 and 4.4. Data were analyzed using the high-throughput HRIBO pipeline and previously undetected adaptations of E. coli to acid stress were found including up-regulation of glycerol catabolism and siderophore production, down-regulation of many membrane proteins and regulation by the transcriptional regulators YdeO, MhpR, IscR, and YdcI. Several examples of differential transcriptional and translational regulation of genes were identified as well as potential novel small open reading frames. These results expand the acid resistance network and provide new insights into the fine-tuned response of E. coli.