Genome-wide Functional Characterization of Escherichia coli Promoters and Sequence Elements Encoding Their Regulation
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ABSTRACT:
ORGANISM(S): Escherichia coli str. K-12 substr. MG1655 Escherichia coli DH5[alpha]
PROVIDER: GSE144621 | GEO | 2020/06/01
REPOSITORIES: GEO
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Project description:Systematic Dissection of Sequence Elements Controlling σ70 Promoters Using a Genomically-Encoded Multiplexed Reporter Assay in E. coli
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Project description:Shimoni2009 - Escherichia Coli SOS
Simple model, involving only the basic components of the circuit, sufficient to explain the peaks in the promoter activities of recA and lexA.
This model is described in the article:
Stochastic analysis of the SOS response in Escherichia coli.
Shimoni Y, Altuvia S, Margalit H, Biham O
PloS one. 2009; 4(5):e5363
Abstract:
BACKGROUND: DNA damage in Escherichia coli evokes a response mechanism called the SOS response. The genetic circuit of this mechanism includes the genes recA and lexA, which regulate each other via a mixed feedback loop involving transcriptional regulation and protein-protein interaction. Under normal conditions, recA is transcriptionally repressed by LexA, which also functions as an auto-repressor. In presence of DNA damage, RecA proteins recognize stalled replication forks and participate in the DNA repair process. Under these conditions, RecA marks LexA for fast degradation. Generally, such mixed feedback loops are known to exhibit either bi-stability or a single steady state. However, when the dynamics of the SOS system following DNA damage was recently studied in single cells, ordered peaks were observed in the promoter activity of both genes (Friedman et al., 2005, PLoS Biol. 3(7):e238). This surprising phenomenon was masked in previous studies of cell populations. Previous attempts to explain these results harnessed additional genes to the system and deployed complex deterministic mathematical models that were only partially successful in explaining the results.
PRINCIPAL FINDINGS: Here we apply stochastic methods, which are better suited for dynamic simulations of single cells. We show that a simple model, involving only the basic components of the circuit, is sufficient to explain the peaks in the promoter activities of recA and lexA. Notably, deterministic simulations of the same model do not produce peaks in the promoter activities.
SIGNIFICANCE: We conclude that the double negative mixed feedback loop with auto-repression accounts for the experimentally observed peaks in the promoter activities. In addition to explaining the experimental results, this result shows that including additional regulations in a mixed feedback loop may dramatically change the dynamic functionality of this regulatory module. Furthermore, our results suggests that stochastic fluctuations strongly affect the qualitative behavior of important regulatory modules even under biologically relevant conditions, thus emphasizing the importance of stochastic analysis of regulatory circuits.
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2005-01-01 | MODEL2937159804 | BioModels
Project description:Escherichia coli strain C is the last of five E. coli strains (C, K12, B, W, Crooks) designated as safe for laboratory purposes whose genome has not been sequenced. We found that E. coli C forms more robust biofilms than the other four laboratory strains. Here we present the complete genomic sequence of this strain in which we utilized high resolution optical mapping to confirm a large inversion in comparison to other strains. DNA sequence comparison revealed the absence of several genes involved in biofilm formation, such as antigen 43, waaSBOJYZUL for LPS synthesis, and cpsB for curli synthesis. The main difference affecting biofilm formation is the presence of an IS3-like insertion sequence in front of the carbon storage regulator csrA gene. This insertion is located 86 bp upstream of the csrA start codon inside the -35 region of P4 promoter and blocks the transcription from the sigma32 and sigma70 promoters P1-P3 located further upstream. Analysis of gene expression profiles in planktonic and biofilm attached cells by the RNAseq method allows better understanding of this regulatory pathway in E. coli.
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Project description:We built and tested a library of 10,898 σ70 promoter variants consisting of all combinations of a set of eight -35 elements, eight -10 elements, three UP elements, eight spacers, and eight backgrounds. Using a Massively Parallel Reporter Assay, we measure expression of all promoters and evaluate how individual promoter elements contribute to expression.
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2020-07-29 | E-MTAB-9036 | biostudies-arrayexpress
Project description:Genome-wide Functional Characterization of Escherichia coli Promoters and Sequence Elements Encoding Their Regulation
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Project description:Bacteria regulate gene expression to adapt to changing environments through transcriptional regulatory networks (TRNs). Although extensively studied, TRNs are not fully characterized since we do not know the identity of all the transcriptional regulators that comprise a TRN. Here, we experimentally evaluate 40 uncharacterized proteins in the model organisms Escherichia coli K-12 MG1655, which were previously predicted to be transcription factors (TFs). First, we used myc-tagging and a multiplexed ChIP-exo assay to characterize genome-wide binding sites for the 40 candidate TFs. Then, we compared those binding sites with those of RNA polymerase and sigma factors to increase the confidence in the measured DNA-binding events. Finally, we used these data to infer potential functional annotation for 13 candidate TFs. As a result, we characterized 570 new binding sites for 34 of 40 candidate TFs. Taken together, the study; 1) expands the number of confirmed TFs from 249 to 278, that is close to the estimated total of about 280 TFs in E. coli K-12 MG1655, 2) it provides a roadmap for functional characterization of the 34 discovered TFs, and 3) it enables the elucidation of the first comprehensive TRN.
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