Corynebacterium glutamicum contains genes for 13 two-component signal transduction systems. In order to test for their essentiality and involvement in the adaptive response to phosphate (P i ) starvation, a set of 12 deletion mutants was constructed. One of the mutants was specifically impaired in its ability to grow under P i limitation, and therefore the genes lacking in this strain were named phoS (encoding the sensor kinase) and phoR (encoding the response regulator). DNA microarray analyses with the C. glutamicum wild type and the ⌬phoRS mutant supported a role for the PhoRS system in the adaptation to P i starvation. In contrast to the wild type, the ⌬phoRS mutant did not induce the known P i starvation-inducible ( psi) genes within 1 hour after a shift from P i excess to P i limitation, except for the pstSCAB operon, which was still partially induced. This indicates an activator function for PhoR and the existence of at least one additional regulator of the pst operon. Primer extension analysis of selected psi genes ( pstS, ugpA, phoR, ushA, and nucH ) confirmed the microarray data and provided evidence for positive autoregulation of the phoRS genes.Phosphorus (P) is an essential nutrient for all cells and required, e.g., for the biosynthesis of nucleotides, DNA, and RNA and in addition for the functional regulation of protein activity by phosphorylation. The common phosphorus source is inorganic phosphate (P i ), and cells have developed mechanisms for the acquisition, assimilation, and storage of phosphate. Under phosphate starvation, many bacteria induce the synthesis of proteins that enable them to use the limiting phosphate resources more efficiently and to make alternative phosphorus sources accessible. The corresponding genes are collectively named P i starvation-inducible genes, or psi genes. The phosphate starvation response, in particular its regulation, has been most carefully studied in Escherichia coli (33) and Bacillus subtilis (11). In both species, two-component signal transduction systems consisting of a histidine kinase and a response regulator play a prominent role.In E. coli, induction of the P i starvation genes is dependent on the PhoR-PhoB two-component system. Under P i limitation, the histidine kinase PhoR phosphorylates the response regulator PhoB, and PhoBϳP in turn activates transcription of at least 31 genes, which form the Pho regulon (33). The genes include the phoBR operon; the pstSCAB-phoU operon, encoding an ABC transporter for high-affinity P i uptake and a regulatory protein; the ugpBAECQ operon, encoding an sn-glycerol 3-phosphate ABC uptake system and glycerophosphoryl diester phosphodiesterase; the phoA-psiF operon, encoding alkaline phosphatase and a protein of unknown function; phoE, encoding an anion-specific porin; phoH, encoding an ATPbinding protein of unknown function; and the phnCDEFGHI JKLMNOP operon, encoding proteins involved in the uptake of phosphonates and their degradation via the C-P lyase pathway. Thus, when P i is scarce, E. coli takes up P i by an ...

Phosphorus is an essential component of macromolecules, like DNA, and central metabolic intermediates, such as sugar phosphates, and bacteria possess enzymes and control mechanisms that provide an optimal supply of phosphorus from the environment. UDP-sugar hydrolases and 5 nucleotidases may play roles in signal transduction, as they do in mammals, in nucleotide salvage, as demonstrated for UshA of Escherichia coli, or in phosphorus metabolism. The Corynebacterium glutamicum gene ushA was found to encode a secreted enzyme which is active as a 5 nucleotidase and a UDP-sugar hydrolase. This enzyme was synthesized and secreted into the medium when C. glutamicum was starved for inorganic phosphate. UshA was required for growth of C. glutamicum on AMP and UDP-glucose as sole sources of phosphorus. Thus, in contrast to UshA from E. coli, C. glutamicum UshA is an important component of the phosphate starvation response of this species and is necessary to access nucleotides and related compounds as sources of phosphorus.

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BackgroundThe pstSCAB operon of Corynebacterium glutamicum, which encodes an ABC transport system for uptake of phosphate (Pi), is induced during the Pi starvation response. The two-component regulatory system PhoRS is involved in this response, but partial Pi starvation induction of pstSCAB in a ΔphoRS mutant indicated the involvement of additional regulator(s). Regulation of pstSCAB also involves the global transcriptional regulator GlxR.ResultsDNA affinity chromatography identified the regulator of acetate metabolism RamB as a protein binding to pstS promoter DNA in vitro. Gel mobility shift assays and mutational analysis of the pstS promoter region revealed that RamB binds to two sites localized at positions −74 to −88 and −9 to +2 with respect to the transcriptional start site of pstSCAB. Reporter gene studies supported the in vivo relevance of both binding sites for activation of pstSCAB by RamB. DNA microarray analysis revealed that expression of many Pi starvation genes reached higher levels during the Pi starvation response on minimal medium with glucose as sole carbon source than in Pi starved acetate-grown C. glutamicum cells.ConclusionsIn C. glutamicum, RamB is involved in expression control of pstSCAB operon. Thus, transcriptional regulation of pstSCAB is complex involving activation by the phosphate-responsive two-component regulatory system PhoSR and the regulators of carbon metabolism GlxR and RamB.

Corynebacterium glutamicum is the major host for the industrial production of amino acids and has become one of the best studied model organisms in microbial biotechnology. Rational strain construction has led to an improvement of producer strains and to a variety of novel producer strains with a broad substrate and product spectrum. A key factor for the success of these approaches is detailed knowledge of transcriptional regulation in C. glutamicum. Here, we present a large compendium of 927 manually curated microarray-based transcriptional profiles for wild-type and engineered strains detecting genome-wide expression changes of the 3,047 annotated genes in response to various environmental conditions or in response to genetic modifications. The replicates within the 927 experiments were combined to 304 microarray sets ordered into six categories that were used for differential gene expression analysis. Hierarchical clustering confirmed that no outliers were present in the sets. The compendium provides a valuable resource for future fundamental and applied research with C. glutamicum and contributes to a systemic understanding of this microbial cell factory. Measurement(s) Gene Expression Analysis Technology Type(s) Two Color Microarray Factor Type(s) WT condition A vs. WT condition B • Plasmid-based gene overexpression in parental strain vs. parental strain with empty vector control • Deletion mutant vs. parental strain Sample Characteristic - Organism Corynebacterium glutamicum Sample Characteristic - Environment laboratory environment Sample Characteristic - Location Germany