The<scp>Crc</scp>protein inhibits the production of polyhydroxyalkanoates in<scp><i>P</i></scp><i>seudomonas putida</i>under balanced carbon/nitrogen growth conditions

Rosa, Ruggero La; Peña, Fernando de la; Prieto, María Axiliadora; Rojo, Fernando

doi:10.1111/1462-2920.12303

Cited by 37 publications

(43 citation statements)

References 65 publications

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“…This stable complex was not detected with the control RNA lacking a CA motif. Similar results were obtained with other RNA oligonucleotides containing known CA motifs, like those present in benR mRNA (Moreno and Rojo, ) or in phaC1 mRNA (La Rosa et al ., ), see Fig. B and C. If Crc was substituted by a similar amount of an unrelated protein such as bovine serum albumin (BSA), the stable complex was not observed (Fig.…”

Section: Resultsmentioning

confidence: 94%

The Crc and Hfq proteins of Pseudomonas putida cooperate in catabolite repression and formation of ribonucleic acid complexes with specific target motifs

Moreno

Hernández-Arranz

Rosa

et al. 2014

Environmental Microbiology

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108

138

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The Crc protein is a global regulator that has a key role in catabolite repression and optimization of metabolism in Pseudomonads. Crc inhibits gene expression post-transcriptionally, preventing translation of mRNAs bearing an AAnAAnAA motif [the catabolite activity (CA) motif] close to the translation start site. Although Crc was initially believed to bind RNA by itself, this idea was recently challenged by results suggesting that a protein co-purifying with Crc, presumably the Hfq protein, could account for the detected RNA-binding activity. Hfq is an abundant protein that has a central role in post-transcriptional gene regulation. Herein, we show that the Pseudomonas putida Hfq protein can recognize the CA motifs of RNAs through its distal face and that Crc facilitates formation of a more stable complex at these targets. Crc was unable to bind RNA in the absence of Hfq. However, pull-down assays showed that Crc and Hfq can form a co-complex with RNA containing a CA motif in vitro. Inactivation of the hfq or the crc gene impaired catabolite repression to a similar extent. We propose that Crc and Hfq cooperate in catabolite repression, probably through forming a stable co-complex with RNAs containing CA motifs to result in inhibition of translation initiation.

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Section: Resultsmentioning

confidence: 94%

The Crc and Hfq proteins of Pseudomonas putida cooperate in catabolite repression and formation of ribonucleic acid complexes with specific target motifs

Moreno

Hernández-Arranz

Rosa

et al. 2014

Environmental Microbiology

Self Cite

108

138

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“…Generally, the action of Crc is limited to the exponential growth phase and it is not observed when the cultures entered into the stationary phase (La Rosa et al 2014). In our study the number of mRNA transcripts of Crc decreased in the stationary phase, but they were still present, most likely reducing mcl-PHAs synthesis by repressing phaC1 translation.…”

Section: Discussionmentioning

confidence: 99%

Medium-chain-length polyhydroxyalkanoates synthesis by Pseudomonas putida KT2440 relA/spoT mutant: bioprocess characterization and transcriptome analysis

et al. 2017

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Pseudomonas putida KT2440 is a model bacteria used commonly for medium-chain-length polyhydroxyalkanoates (mcl-PHAs) production using various substrates. However, despite many studies conducted on P. putida KT2440 strain, the molecular mechanisms of leading to mcl-PHAs synthesis in reaction to environmental stimuli are still not clear. The rearrangement of the metabolism in response to environmental stress could be controlled by stringent response that modulates the transcription of many genes in order to promote survival under nutritional deprivation conditions. Therefore, in this work we investigated the relation between mcl-PHAs synthesis and stringent response. For this study, a relA/spoT mutant of P. putida KT2440, unable to induce the stringent response, was used. Additionally, the transcriptome of this mutant was analyzed using RNA-seq in order to examine rearrangements of the metabolism during cultivation. The results show that the relA/spoT mutant of P. putida KT2440 is able to accumulate mcl-PHAs in both optimal and nitrogen limiting conditions. Nitrogen starvation did not change the efficiency of mcl-PHAs synthesis in this mutant. The transition from exponential growth to stationary phase caused significant upregulation of genes involved in transport system and nitrogen metabolism. Transcriptional regulators, including rpoS, rpoN and rpoD, did not show changes in transcript abundance when entering the stationary phase, suggesting their limited role in mcl-PHAs accumulation during stationary phase.Electronic supplementary materialThe online version of this article (doi:10.1186/s13568-017-0396-z) contains supplementary material, which is available to authorized users.

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“…52,[66][67][68][69][70] To ascertain if the bacteria of interest in this study accumulate these intracellular carbon-storage products from APL catabolism, such as PHAs and fatty acids, Nile red staining and flow cytometry were conducted for the bacterial strains in the nitrogen-limiting conditions each day of the cultivation as shown in Fig. 52,[66][67][68][69][70] To ascertain if the bacteria of interest in this study accumulate these intracellular carbon-storage products from APL catabolism, such as PHAs and fatty acids, Nile red staining and flow cytometry were conducted for the bacterial strains in the nitrogen-limiting conditions each day of the cultivation as shown in Fig.…”

Section: Generation Of Intracellular Co-products During Bacterial Gromentioning

confidence: 99%

Towards lignin consolidated bioprocessing: simultaneous lignin depolymerization and product generation by bacteria

et al. 2015

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Lignin represents an untapped resource in lignocellulosic biomass, primarily due to its recalcitrance to depolymerization and its intrinsic heterogeneity. In Nature, microorganisms have evolved mechanisms to both depolymerize lignin using extracellular oxidative enzymes and to uptake the aromatic species generated during depolymerization for carbon and energy sources. The ability of microbes to conduct both of these processes simultaneously could enable a Consolidated Bioprocessing concept to be applied to lignin, similar to what is done today with polysaccharide conversion to ethanol via ethanologenic, cellulolytic microbes. To that end, here we examine the ability of 14 bacteria to secrete ligninolytic enzymes, depolymerize lignin, uptake aromatic and other compounds present in a biomass-derived, lignin-enriched stream, and, under nitrogen-limiting conditions, accumulate intracellular carbon storage compounds that can be used as fuel, chemical, or material precursors. In shake flask conditions using a substrate produced during alkaline pretreatment, we demonstrate that up to nearly 30% of the initial lignin can be depolymerized and catabolized by a subset of bacteria. In particular, Amycolatopsis sp., two Pseudomonas putida strains, Acinetobacter ADP1, and Rhodococcus jostii are able to depolymerize high molecular weight lignin species and catabolize a significant portion of the low molecular weight aromatics, thus representing good starting hosts for metabolic engineering. This study also provides a comprehensive set of experimental tools to simultaneously study lignin depolymerization and aromatic catabolism in bacteria, and provides a foundation towards the concept of Lignin Consolidated Bioprocessing, which may eventually be an important route for biological lignin valorization. Electronic Supplementary Information (ESI) available: Additional resultsdescribing the HPAE-PAD findings, growth curves of the bacteria over the 7-day experiments, the full set of ligninolytic enzyme profiles, and genomic analyses for ligninolytic enzymes and aromatic catabolic enzymes. See

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TheCrcprotein inhibits the production of polyhydroxyalkanoates inPseudomonas putidaunder balanced carbon/nitrogen growth conditions

Cited by 37 publications

References 65 publications

The Crc and Hfq proteins of Pseudomonas putida cooperate in catabolite repression and formation of ribonucleic acid complexes with specific target motifs

The Crc and Hfq proteins of Pseudomonas putida cooperate in catabolite repression and formation of ribonucleic acid complexes with specific target motifs

Medium-chain-length polyhydroxyalkanoates synthesis by Pseudomonas putida KT2440 relA/spoT mutant: bioprocess characterization and transcriptome analysis

Towards lignin consolidated bioprocessing: simultaneous lignin depolymerization and product generation by bacteria

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