Revisiting the mechanism of activation of cyclic AMP receptor protein (CRP) by cAMP in <i>Escherichia coli</i>: Lessons from a subunit‐crosslinked form of CRP

Saha, Abinit; Mukhopadhyay, Jayanta; Datta, Aritreyee; Parrack, Pradeep

doi:10.1016/j.febslet.2014.12.021

Cited by 10 publications

(9 citation statements)

References 24 publications

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“…Interestingly, the expressed mRNA and protein level of both crp or cra does not change significantly during growth on glucose, fructose, or acetate ( Supplementary Figure S10 , Supplementary Figure S11 ). Therefore, the regulatory activity of CRP could be strongly dependent on the concentration of its effector molecule, cAMP, as suggested in the previous study with in vitro experiment ( 52 ). The intracellular concentration of cAMP is lowest on glucose, and higher on fructose.…”

Section: Resultsmentioning

confidence: 78%

Systems assessment of transcriptional regulation on central carbon metabolism by Cra and CRP

Kim

Seo

Gao

et al. 2018

Nucleic Acids Research

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Two major transcriptional regulators of carbon metabolism in bacteria are Cra and CRP. CRP is considered to be the main mediator of catabolite repression. Unlike for CRP, in vivo DNA binding information of Cra is scarce. Here we generate and integrate ChIP-exo and RNA-seq data to identify 39 binding sites for Cra and 97 regulon genes that are regulated by Cra in Escherichia coli. An integrated metabolic-regulatory network was formed by including experimentally-derived regulatory information and a genome-scale metabolic network reconstruction. Applying analysis methods of systems biology to this integrated network showed that Cra enables optimal bacterial growth on poor carbon sources by redirecting and repressing glycolysis flux, by activating the glyoxylate shunt pathway, and by activating the respiratory pathway. In these regulatory mechanisms, the overriding regulatory activity of Cra over CRP is fundamental. Thus, elucidation of interacting transcriptional regulation of core carbon metabolism in bacteria by two key transcription factors was possible by combining genome-wide experimental measurement and simulation with a genome-scale metabolic model.

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Section: Resultsmentioning

confidence: 78%

Systems assessment of transcriptional regulation on central carbon metabolism by Cra and CRP

Kim

Seo

Gao

et al. 2018

Nucleic Acids Research

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“…The evidence that both mutants, Δ crp (Q-type) and Δ crp (R-type), shared some common target genes (13.3% in the up-regulated genes, 16% in the down-regulated genes, Figure 4 ) during the exponential phase was somewhat unexpected. A first, simple interpretation of this phenomenon could suggest that transcription of these sets of genes does not depend on the acetylation stage of CRP, but rather on the presence of several conformer complexes between c-AMP and CRP (Heyduk and Lee, 1989 ; Mukhopadhyay et al, 1999 ; Tworzydło et al, 2005 ; Tutar, 2008 ; Saha et al, 2015 ).…”

Section: Resultsmentioning

confidence: 99%

“…In addition, the crp gene has two binding sites for the CRP protein, a stimulatory one located upstream of the promoter, and an inhibitory one located downstream of the promoter (Ishizuka et al, 1994 ). CRP is not really what is bound, but rather the cAMP-CRP complex (Saha et al, 2015 ) (see Figure 6 ).…”

Section: Resultsmentioning

confidence: 99%

Influence of Glucose Availability and CRP Acetylation on the Genome-Wide Transcriptional Response of Escherichia coli: Assessment by an Optimized Factorial Microarray Analysis

Guebel¹,

Torres

2018

Front. Microbiol.

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Background: While in eukaryotes acetylation/deacetylation regulation exerts multiple pleiotropic effects, in Escherichia coli it seems to be more limited and less known. Hence, we aimed to progress in the characterization of this regulation by dealing with three convergent aspects: the effector enzymes involved, the master regulator CRP, and the dependence on glucose availability.Methods: The transcriptional response of E. coli BW25113 was analyzed across 14 relevant scenarios. These conditions arise when the wild type and four isogenic mutants (defective in deacetylase CobB, defective in N(ε)-lysine acetyl transferase PatZ, Q- and R-type mutants of protein CRP) are studied under three levels of glucose availability (glucose-limited chemostat and glucose-excess or glucose-exhausted in batch culture). The Q-type emulates a permanent stage of CRPacetylated, whereas the R-type emulates a permanent stage of CRPdeacetylated. The data were analyzed by an optimized factorial microarray method (Q-GDEMAR).Results: (a) By analyzing one mutant against the other, we were able to unravel the true genes that participate in the interaction between ΔcobB/ΔpatZ mutations and glucose availability; (b) Increasing stages of glucose limitation appear to be associated with the up-regulation of specific sets of target genes rather than with the loss of genes present when glucose is in excess; (c) Both CRPdeacetylated and CRPacetylated produce extensive changes in specific subsets of genes, but their number and identity depend on the glucose availability; (d) In other sub-sets of genes, the transcriptional effect of CRP seems to be independent of its acetylation or deacetylation; (e) Some specific ontology functions can be associated with each of the different sets of genes detected herein.Conclusions: CRP cannot be thought of only as an effector of catabolite repression, because it acts along all the glucose conditions tested (excess, limited, and exhausted), exerting both positive and negative effects through different sets of genes. Acetylation of CRP does not seem to be a binary form of regulation, as there is not a univocal relationship between its activation/inhibitory effect and its acetylation/deacetylation stage. All the combinatorial possibilities are observed. During the exponential growth phase, CRP also exerts a very significant transcriptional effect, mainly on flagellar assembly and chemotaxis (FDR = 7.2 × 10−44).

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“…These include reorientation of the DNA-binding domain and stabilization of the backbone helix, but the two different structures do not agree on the orientation of the C-terminal domain. This observation, and the limited number of published apo CRP structures, suggests that apoCRP may be unstable due to flexibility of the C-terminal domain [44].…”

Section: Introductionmentioning

confidence: 99%

Mutations in the Global Transcription Factor CRP/CAP: Insights from Experimental Evolution and Deep Sequencing

Frendorf

Lauritsen

Sekowska

et al. 2019

Computational and Structural Biotechnology Journal

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The Escherichia coli cyclic AMP receptor protein (CRP or catabolite activator protein, CAP) provides a textbook example of bacterial transcriptional regulation and is one of the best studied transcription factors in biology. For almost five decades a large number of mutants, evolved in vivo or engineered in vitro, have shed light on the molecular structure and mechanism of CRP. Here, we review previous work, providing an overview of studies describing the isolation of CRP mutants. Furthermore, we present new data on deep sequencing of different bacterial populations that have evolved under selective pressure that strongly favors mutations in the crp locus. Our new approach identifies more than 100 new CRP mutations and paves the way for a deeper understanding of this fascinating bacterial master regulator.

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Revisiting the mechanism of activation of cyclic AMP receptor protein (CRP) by cAMP in Escherichia coli: Lessons from a subunit‐crosslinked form of CRP

Cited by 10 publications

References 24 publications

Systems assessment of transcriptional regulation on central carbon metabolism by Cra and CRP

Systems assessment of transcriptional regulation on central carbon metabolism by Cra and CRP

Influence of Glucose Availability and CRP Acetylation on the Genome-Wide Transcriptional Response of Escherichia coli: Assessment by an Optimized Factorial Microarray Analysis

Mutations in the Global Transcription Factor CRP/CAP: Insights from Experimental Evolution and Deep Sequencing

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