title: Ethambutol, a cell wall inhibitor of Mycobacterium tuberculosis, elicits L-glutamate efflux of Corynebacterium glutamicum.
abstract: Corynebacterium glutamicum is used for the large-scale production of L-glutamate, but the efflux of this amino acid is poorly understood. This study shows that addition of ethambutol (EMB) to growing cultures of C. glutamicum causes L-glutamate efflux at rates of up to 15 nmol min(-1) (mg dry wt)(-1), whereas in the absence of EMB, no efflux occurs. EMB is used for the treatment of Mycobacterium tuberculosis, and at a molecular level it targets a series of arabinosyltransferases (EmbCAB). The single arabinosyltransferase-encoding emb gene of C. glutamicum was placed under the control of a Tet repressor (TetR). Experiments with this strain, as well as with an emb-overexpressing strain, coupled with biochemical analyses showed that: (i) emb expression was correlated with L-glutamate efflux, (ii) emb overexpression increased EMB resistance, (iii) EMB caused less arabinan deposition in cell wall arabinogalactan, and (iv) EMB caused a reduced content of cell-wall-bound mycolic acids. Thus EMB addition resulted in a marked disordering of the cell envelope, which was also discernible by examining cellular morphology. In order to further characterize the cellular response to EMB addition, genome-wide expression profiling was performed using DNA microarrays. This identified 76 differentially expressed genes, with 18 of them upregulated more than eightfold. Among these were the cell-wall-related genes ftsE and mepA (encoding a secreted metalloprotease); however, genes of central metabolism were largely absent. Given that an altered lipid composition of the plasma membrane of C. glutamicum can result in L-glutamate efflux, we speculate that major structural alterations of the cell envelope are transmitted to the membrane, which in turn activates an export system, perhaps via increased membrane tension.
authors.affiliation: Institute for Biotechnology, Research Centre Jülich, D-52425 Jülich, Germany.
authors.forename: Eva
authors.initials: E
authors.lastname: Radmacher
authors.affiliation: Institute for Biotechnology, Research Centre Jülich, D-52425 Jülich, Germany.
authors.forename: Kathrin C
authors.initials: KC
authors.lastname: Stansen
authors.affiliation: School of Biosciences, University of Birmingham, Birmingham B15 2TT, UK.
authors.forename: Gurdyal S
authors.initials: GS
authors.lastname: Besra
authors.affiliation: School of Biosciences, University of Birmingham, Birmingham B15 2TT, UK.
authors.forename: Luke J
authors.initials: LJ
authors.lastname: Alderwick
authors.affiliation: School of Biosciences, University of Birmingham, Birmingham B15 2TT, UK.
authors.forename: William N
authors.initials: WN
authors.lastname: Maughan
authors.affiliation: Pathology, University Hospital, D-52074 Aachen, Germany.
authors.forename: Günter
authors.initials: G
authors.lastname: Hollweg
authors.affiliation: Institute for Biotechnology, Research Centre Jülich, D-52425 Jülich, Germany.
authors.forename: Hermann
authors.initials: H
authors.lastname: Sahm
authors.affiliation: Institute for Biotechnology, Research Centre Jülich, D-52425 Jülich, Germany.
authors.forename: Volker F
authors.initials: VF
authors.lastname: Wendisch
authors.affiliation: Institute for Biotechnology, Research Centre Jülich, D-52425 Jülich, Germany.
authors.forename: Lothar
authors.initials: L
authors.lastname: Eggeling
mesh_terms: D000995:Antitubercular Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002473:Cell Wall / Q000187:drug effects; D048230:Corynebacterium glutamicum / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D024881:Drug Resistance, Bacterial; D004977:Ethambutol / Q000494:pharmacology*; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D016680:Genome, Bacterial; D005971:Glutamates / Q000378:metabolism*; D008826:Microbial Sensitivity Tests; D009169:Mycobacterium tuberculosis / Q000187:drug effects; D009171:Mycolic Acids / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D010430:Pentosyltransferases / Q000235:genetics / Q000378:metabolism; D011134:Polysaccharides / Q000378:metabolism
chemical_list: D000995:Antitubercular Agents; D001426:Bacterial Proteins; D005971:Glutamates; D009171:Mycolic Acids; D011134:Polysaccharides; C030080:araban; D004977:Ethambutol; D010430:Pentosyltransferases; C101967:arabinosyltransferase