Contact : Marie-Joëlle VIROLLE
Associate Scientist - Principal Team Investigator
Phone : +331 69 15 46 42
Building 400, Room 110
Marie-Joëlle VIROLLE, Associate Scientist, CNRS
Catherine ESNAULT, Lecturer, University of Paris VI
Fabien COZE, Ph. D. student
Thierry DULERMO, Post-Doc
The group of « Métabolisme Energétique des Streptomyces » (MES) studies the regulatory switch governing the transition between primary and secondary metabolism, in the bacteria of great industrial interest, Streptomyces.
The Streptomyces are filamentous Gram + bacteria living in the superficial layers of the soil where they contribute to the degradation of organic debris. These bacteria are of industrial interest since they produce numerous metabolites, diverse in chemical structures and biological activities, that are often useful to human health (antibiotics, anti-cancer drugs, immunosuppressive-drugs, anti-inflammatory drugs etc…) or agriculture (herbicide, fungicide, insecticide etc…). These compounds, usually called « secondary metabolites », are synthesised during the periods of slow or no growth (weak anabolism). Their synthesis is thought to be triggered by some nutritional limitations, a nutritional limitation in phosphate being the most efficient trigger. These “secondary metabolites” are synthesised from precursors originating from the primary metabolism (sugars, amino acids, nucleosides, short fatty acids etc…). These building blocks are modified and combined by large multi-functional enzymatic complexes (PKS, NRPS…) encoded by groups of genes constituting biosynthetic clusters. The expression of the genes of these clusters is coordinated by specific, positive or negative, regulators linked to the pathways. However, the regulatory cascade that goes from the « sensing » of a nutritional limitation in phosphate to the up or down regulation of the positive or negative specific regulators of the biosynthetic pathways is still poorly understood.
The group MES characterised a gene (ppk) and an operon (phoR/phoP), both linked to Pi metabolism, whose interruption leads to an important increase in antibiotic biosynthesis in the weak producer Streptomyces lividans (3). Furthermore, the group MES assessed the role of the polyphosphates (polymer of phosphate linked by high energy phosphoanhydride bonds) in the regulation of antibiotics biosynthesis. Physiological studies as well as some partial transcriptomic and proteomic analysis of the ppk and the phoP mutant strains of S. lividans led the group MES to propose a novel model of regulation of antibiotic biosynthesis that has high explicative and predictive values (see below).
The ppk gene encodes a protein bearing strong similarities with polyphosphate kinases of various origins (3).
The group MES demonstrated that, in Streptomyces, this enzyme catalyses the regeneration of ATP from ADP and polyphosphates. Ppk thus plays an important role in the energetic metabolism of the cell and the ppk mutant is predicted to have a deficit in its energetic charge compared to the wild type strain. In order to compensate for this energetic deficit, a strong activation of the central metabolic pathways (glycolysis, Krebs cycle coupled to the respiratory chain) would be triggered in the ppk mutant strain. This activation of central metabolism would lead to the synthesis of ATP, of reduced co-factors (NADH/NADPH) and of carbon skeletons. Since this activation occurs in the periods of weak anabolism, these molecules are likely to be produced in large excess relatively to the cell needs for maintenance and survival. In consequence these molecules are likely to be re-directed toward antibiotic biosynthesis (1). This activation, in the process to be confirmed by transcriptomic and proteomic approaches, requires an enhanced supply of carbon, and phosphate, fuels of the metabolism, and leads to an increase of the internal oxidative stress. This increase is thought to be due to a saturation of the respiratory chain by the too abundant reduced co-factors (NADH, NADPH) produced by the activation of the metabolism. This saturation of the respiratory chain would lead to electrons leakage and thus to the generation of free radicals (ROS, RNS etc…) (2).
The two components system phoR (sensory kinase) / phoP (response regulator) controls the expression of genes of the pho regulon that includes genes involved in the supply of the phosphate (ex: genes encoding the high affinity phosphate transporter or various phosphatases etc…) or in ATP regeneration (ex: ppk). Genes of the pho regulon thus play an important role in the energetic metabolism of the cell, in condition of phosphate limitation. The phoP mutant likely triggers similar strategies than the ppk mutant to generate the necessary ATP, in condition of Pi limitation. However, since this mutant is unable to provide sufficient free phosphate to sustain the activation of the metabolism, this activation and thus precursors supply and antibiotic production cannot last very long in this mutant that lyses precociously (2).
Seghezzi N, Amar P, Koebmann B, Jensen PR and Virolle MJ, (2011), The construction of a library of synthetic promoters revealed some specific features of strong Streptomyces promoters, Appl Microbiol Biotechnol
Xu D, Seghezzi N, Esnault C and Virolle MJ, (2010), The Over-expression of a Transcriptional Regulator of the TetR family Represses Antibiotic production and Sporulation in Streptomyces coelicolor, Appl Environ Microbiol, Epub ahead of print
Hamdali H, Virolle MJ, von Jan M, Sproer C, Klenk HP and Ouhdouch Y, (2010), Streptomyces youssoufiensis sp. nov., isolated from a Moroccan phosphate mine, Int J Syst Evol Microbiol, Epub ahead of print
Xu D, Seghezzi N, Esnault C and Virolle MJ, (2010), Repression of antibiotic production and sporulation in Streptomyces coelicolor by overexpression of a TetR family transcriptional regulator, Appl Environ Microbiol, 76(23):7741-53
Ghorbel S, Smirnov A, Chouayekh H, Sperandio B, Esnault C, Kormanec J and Virolle MJ, (2006), Regulation of ppk Expression and In Vivo Function of Ppk in Streptomyces lividans TK24, J. Bacteriol. 188 (17):6269-76
Ghorbel S, Kormanec J, Artus A and Virolle MJ, (2006), Transcriptional studies and regulatory interactions between the phoR-phoP operon and the phoU, mtpA, and ppk genes of Streptomyces lividans TK24, J. Bacteriol. 188 (2):677-86
Chouayekh H and Virolle MJ, (2002), The polyphosphate kinase is involved in the phosphate control of actinorhodin production in Streptomyces lividans, Mol. Microbiol. 43(4):919-30
Chouayekh H, Nothaft H, Delaunay S, Linder M, Payrastre B, Seghezzi N, Titgemeyer F and Virolle MJ, (2007), Phosphoinositide are involved in the control of the growth resumption that follows the transition phase in Streptomyces lividans, in the presence of glucose, J. Bacteriol. (189) (3) : 741–49
Gagnat J, Chouayekh H, Gerbaud C, Francou F, Virolle MJ, (1999), Disruption of sblA in Streptomyces lividans permits expression of a heterologous alpha-amylase gene in the presence of glucose, Microbiology 145 ( Pt 9):2303-12