The <i>Pseudomonas putida</i> Crc global regulator controls the hierarchical assimilation of amino acids in a complete medium: Evidence from proteomic and genomic analyses

Moreno, Renata; Martínez-Gomáriz, Montserrat; Yuste, Luís; Gil, Concha; Rojo, Fernando

doi:10.1002/pmic.200800918

Cited by 103 publications

(162 citation statements)

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“…The mechanism through which Crp and CyoB enable growth on BCAA, surprisingly, may also involve the Crc regulator. This is because the Crc global regulator of P. putida KT2440 has been previously shown to repress a number of enzymes involved in BCAA metabolism (34). Hence, we postulate that while Crc may exert inhibitory control over BCAA metabolism, the positive influence of Crp and CyoB trumps this inhibition.…”

Section: Resultsmentioning

confidence: 88%

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Global Regulation of Food Supply by P seudomonas p utida DOT-T1E

et al. 2010

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Pseudomonas putida DOT-T1E was used as a model to develop a "phenomics" platform to investigate the ability of P. putida to grow using different carbon, nitrogen, and sulfur sources and in the presence of stress molecules. Results for growth of wild-type DOT-T1E on 90 different carbon sources revealed the existence of a number of previously uncharted catabolic pathways for compounds such as salicylate, quinate, phenylethanol, gallate, and hexanoate, among others. Subsequent screening on the subset of compounds on which wild-type DOT-TIE could grow with four knockout strains in the global regulatory genes ⌬crc, ⌬crp, ⌬cyoB, and ⌬ptsN allowed analysis of the global response to nutrient supply and stress. The data revealed that most global regulator mutants could grow in a wide variety of substrates, indicating that metabolic fluxes are physiologically balanced. It was found that the Crc mutant did not differ much from the wild-type regarding the use of carbon sources. However, certain pathways are under the preferential control of one global regulator, i.e., metabolism of succinate and D-fructose is influenced by CyoB, and L-arginine is influenced by PtsN. Other pathways can be influenced by more than one global regulator; i.e., L-valine catabolism can be influenced by CyoB and Crp (cyclic AMP receptor protein) while phenylethylamine is affected by Crp, CyoB, and PtsN. These results emphasize the cross talk required in order to ensure proper growth and survival. With respect to N sources, DOT-T1E can use a wide variety of inorganic and organic nitrogen sources. As with the carbon sources, more than one global regulator affected growth with some nitrogen sources; for instance, growth with nucleotides, dipeptides, D-amino acids, and ethanolamine is influenced by Crp, CyoB, and PtsN. A surprising finding was that the Crp mutant was unable to flourish on ammonium. Results for assayed sulfur sources revealed that CyoB controls multiple points in methionine/cysteine catabolism while PtsN and Crc are needed for N-acetyl-L-cysteamine utilization. Growth of global regulator mutants was also influenced by stressors of different types (antibiotics, oxidative agents, and metals). Overall and in combination with results for growth in the presence of various stressors, these phenomics assays provide multifaceted insights into the complex decision-making process involved in nutrient supply, optimization, and survival.

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Section: Resultsmentioning

confidence: 88%

“…3). A recent publication from Fernando Rojo's group included an extensive proteomic and transcriptomic analysis of a P. putida KT2440 ⌬crc mutant strain that had been grown in rich medium (34). It was found that up to 134 genes were differentially expressed in the absence of Crc, with 80% increased in expression and 20% show-FIG.…”

Section: Resultsmentioning

confidence: 99%

Global Regulation of Food Supply by P seudomonas p utida DOT-T1E

et al. 2010

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“…The final role of this regulator is to optimize metabolism, improving bacterial fitness (7). Crc is an RNA-binding protein that recognizes specific sites located at the translation initiation regions of target mRNAs, inhibiting translation (30 -32).…”

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confidence: 99%

“…In Pseudomonas, catabolite repression is not well understood. Available evidence shows that in this bacterial group, certain organic acids and amino acids are the most strongly preferred compounds, whereas hydrocarbons are less preferred; the preference for glucose lies somewhere in between (5)(6)(7). For this reason, the potential of P. putida to degrade aromatic hydrocarbons is frequently compromised by CCR when in addition to the aromatic substrates, other more readily degradable compounds are present.…”

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confidence: 99%

The Crc Global Regulator Inhibits the Pseudomonas putida pWW0 Toluene/Xylene Assimilation Pathway by Repressing the Translation of Regulatory and Structural Genes

Moreno

Fonseca²,

Rojo

2010

Journal of Biological Chemistry

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In Pseudomonas putida, the expression of the pWW0 plasmid genes for the toluene/xylene assimilation pathway (the TOL pathway) is subject to complex regulation in response to environmental and physiological signals. This includes strong inhibition via catabolite repression, elicited by the carbon sources that the cells prefer to hydrocarbons. The Crc protein, a global regulator that controls carbon flow in pseudomonads, has an important role in this inhibition. Crc is a translational repressor that regulates the TOL genes, but how it does this has remained unknown. This study reports that Crc binds to sites located at the translation initiation regions of the mRNAs coding for XylR and XylS, two specific transcription activators of the TOL genes. Unexpectedly, eight additional Crc binding sites were found overlapping the translation initiation sites of genes coding for several enzymes of the pathway, all encoded within two polycistronic mRNAs. Evidence is provided supporting the idea that these sites are functional. This implies that Crc can differentially modulate the expression of particular genes within polycistronic mRNAs. It is proposed that Crc controls TOL genes in two ways. First, Crc inhibits the translation of the XylR and XylS regulators, thereby reducing the transcription of all TOL pathway genes. Second, Crc inhibits the translation of specific structural genes of the pathway, acting mainly on proteins involved in the first steps of toluene assimilation. This ensures a rapid inhibitory response that reduces the expression of the toluene/ xylene degradation proteins when preferred carbon sources become available.

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“…One strategy to this end could be the use of a crc mutant of P. putida to allow efficient translation of XylR during exponential phase. However, as Crc is a global regulator affecting many cellular processes (Moreno et al, 2009;Linares et al, 2010), we adopted instead an experimental protocol to increase XylR expression in a controllable way with minimum interference with cell metabolism and state. To this end, we engineered a mini-Tn7 transposon carrying an expression cassette LacI q -Ptrc → xylR (Perez-Martin and de Lorenzo, 1996) and inserted it into the chromosome of P. putida KT2440 strain bearing in its chromosome a transcriptional Pu-GFP fusion ( Figure 6 and Materials and methods).…”

Section: Overproduction Of Xylr Defeats Pu Bimodality In Vivomentioning

confidence: 99%

Transcription factor levels enable metabolic diversification of single cells of environmental bacteria

2015

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Transcriptional noise is a necessary consequence of the molecular events that drive gene expression in prokaryotes. However, some environmental microorganisms that inhabit polluted sites, for example, the m-xylene degrading soil bacterium Pseudomonas putida mt-2 seem to have co-opted evolutionarily such a noise for deploying a metabolic diversification strategy that allows a cautious exploration of new chemical landscapes. We have examined this phenomenon under the light of deterministic and stochastic models for activation of the main promoter of the master m-xylene responsive promoter of the system (Pu) by its cognate transcriptional factor (XylR). These analyses consider the role of co-factors for Pu activation and determinants of xylR mRNA translation. The model traces the onset and eventual disappearance of the bimodal distribution of Pu activity along time to the growth-phase dependent abundance of XylR itself, that is, very low in exponentially growing cells and high in stationary. This tenet was validated by examining the behaviour of a Pu-GFP fusion in a P. putida strain in which xylR expression was engineered under the control of an IPTG-inducible system. This work shows how a relatively simple regulatory scenario (for example, growth-phase dependent expression of a limiting transcription factor) originates a regime of phenotypic diversity likely to be advantageous in competitive environmental settings.

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The Pseudomonas putida Crc global regulator controls the hierarchical assimilation of amino acids in a complete medium: Evidence from proteomic and genomic analyses

Cited by 103 publications

References 65 publications

Global Regulation of Food Supply by P seudomonas p utida DOT-T1E

Global Regulation of Food Supply by P seudomonas p utida DOT-T1E

The Crc Global Regulator Inhibits the Pseudomonas putida pWW0 Toluene/Xylene Assimilation Pathway by Repressing the Translation of Regulatory and Structural Genes

Transcription factor levels enable metabolic diversification of single cells of environmental bacteria

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