The Carbon Storage Regulator (Csr) System Exerts a Nutrient-Specific Control over Central Metabolism in Escherichia coli Strain Nissle 1917 Olga Revelles 1,2,3. , Pierre Millard 1,2,3. , Jean-Philippe Nougayre `de 4,5,6,7 , Ulrich Dobrindt 8 , Eric Oswald 4,5,6,7 , Fabien Le ´ tisse 1,2,3 , Jean-Charles Portais 1,2,3 * 1 Laboratoire d’Inge ´nierie des Syste `mes Biologiques et des Proce ´de ´s, LISBP, Universite ´ de Toulouse, INSA, UPS, INP, Toulouse, France, 2 Laboratoire Inge ´nierie des Syste `mes Biologiques et des Proce ´de ´s, INRA UMR792, Toulouse, France, 3 UMR5504, CNRS, Toulouse, France, 4 USC1360, INRA, Toulouse, France, 5 U1043, Inserm, Toulouse, France, 6 UMR5282, CNRS, Toulouse, France, 7 Centre de Physiopathologie de Toulouse Purpan, Universite ´ de Toulouse, UPS, Toulouse, France, 8 Institute of Hygiene, University of Mu ¨ nster, Mu ¨ nster, Germany Abstract The role of the post-transcriptional carbon storage regulator (Csr) system in nutrient utilization and in the control of the central metabolism in E. coli reference commensal strain Nissle 1917 was investigated. Analysis of the growth capabilities of mutants altered for various components of the Csr system (csrA51, csrB, csrC and csrD mutations) showed that only the protein CsrA - the key component of the system - exerts a marked role in carbon nutrition. Attenuation of CsrA activity in the csrA51 mutant affects the growth efficiency on a broad range of physiologically relevant carbon sources, including compounds utilized by the Entner-Doudoroff (ED) pathway. Detailed investigations of the metabolomes and fluxomes of mutants and wild-type cells grown on carbon sources representative of glycolysis and of the ED pathway (glucose and gluconate, respectively), revealed significant re-adjusting of central carbon metabolism for both compounds in the csrA51 mutant. However, the metabolic re-adjusting observed on gluconate was strikingly different from that observed on glucose, indicating a nutrient-specific control of metabolism by the Csr system. Citation: Revelles O, Millard P, Nougayre `de J-P, Dobrindt U, Oswald E, et al. (2013) The Carbon Storage Regulator (Csr) System Exerts a Nutrient-Specific Control over Central Metabolism in Escherichia coli Strain Nissle 1917. PLoS ONE 8(6): e66386. doi:10.1371/journal.pone.0066386 Editor: Ching-Hong Yang, University of Wisconsin-Milwaukee, United States of America Received February 7, 2013; Accepted May 5, 2013; Published June 20, 2013 Copyright: ß 2013 Revelles et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: The postdoctoral and PhD fellowships of OR and PM, respectively, were funded by the Institut National de la Recherche Agronomique [PM: Progran Young Researcher]. This work has benefited from the continuous support of the Re ´gion Midi-Pyre ´ne ´es, the European Regional Development Fund, the French Ministry for Higher Education & Research, and SICOVAL, and the Institut National de la Recherche Agronomique. UD was supported by the German Research Foundation [SFB1009,TP B05]. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: portais@insa-toulouse.fr . These authors contributed equally to this work. Introduction Escherichia coli is a normal inhabitant of the intestine and the predominant facultative anaerobe in the gastrointestinal tract of mammals [1]. The intestine is a highly complex and changing environment in which E. coli experiences constantly shifting growth conditions. Recent findings indicate that the ability to compete for carbon nutrition is a critical factor for gut colonization, and is part of the arsenal of strategies employed by pathogenic E. coli strains to outcompete the gut microbiotia [2,3]. Colonization is mainly related to the utilization of sugars and sugar derivatives resulting from the degradation of mucus and of dietary fibers [4,5]. Glycolytic pathways such as the Embden-Meyerhof- Parnas (EMP) and Entner-Doudoroff (ED) pathways play an important role in colonization [4,6,7,8]. Persistence of E. coli in the gut is supported by less favorable substrates, including both sugars and non-sugar compounds such as the small organic acids resulting from the degradation of mucus by anaerobes of the microflora [4,5]. The use of the latter compounds requires activation of gluconeogenic pathways, and efficient switching between glycolytic and gluconeogenic carbon sources is likely to be a major feature of successful adaptation to life in the intestine [9]. To cope with the changing environment in the intestine, E. coli has developed a variety of adaptation mechanisms. Highly sophisticated global regulatory networks coordinate physiological and metabolic responses by controlling the functional expression of relevant sets of genes. The carbon storage regulator (Csr) system [10] is a post-transcriptional regulation system that controls a broad range of physiological adaptative mechanisms (including formation of biofilm, motility and virulence) and is a global regulator of central metabolism [11]. The Csr system has four molecular components (Figure 1). The main component is the post-transcriptional regulator CsrA, a protein that influences both translation and degradation of different mRNA targets [12]. CsrA is negatively regulated by two small non-coding RNAs, CsrB and CsrC, that antagonize CsrA activity by sequestering the protein [13,14]. Lastly, the CsrD protein positively controls CsrA activity by driving the RNAs CsrB and CsrC to RNAse E degradation [15]. The molecular targets of the Csr system include a significant number of metabolic components. Indeed, CsrA is able to interact with most mRNAs of central metabolism enzymes [16], though the actual result of these interactions in terms of mRNA stability and translation is not known for most targets. Biochemical PLOS ONE | www.plosone.org 1 June 2013 | Volume 8 | Issue 6 | e66386