Ethambutol, a cell wall inhibitor of Mycobacterium tuberculosis, elicits L-glutamate efflux of Corynebacterium glutamicum.
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.
Radmacher
Eva
E
Institute for Biotechnology, Research Centre Jülich, D-52425 Jülich, Germany.
Stansen
Kathrin C
KC
Institute for Biotechnology, Research Centre Jülich, D-52425 Jülich, Germany.
Besra
Gurdyal S
GS
School of Biosciences, University of Birmingham, Birmingham B15 2TT, UK.
Alderwick
Luke J
LJ
School of Biosciences, University of Birmingham, Birmingham B15 2TT, UK.
Maughan
William N
WN
School of Biosciences, University of Birmingham, Birmingham B15 2TT, UK.
Hollweg
Günter
G
Pathology, University Hospital, D-52074 Aachen, Germany.
Sahm
Hermann
H
Institute for Biotechnology, Research Centre Jülich, D-52425 Jülich, Germany.
Wendisch
Volker F
VF
Institute for Biotechnology, Research Centre Jülich, D-52425 Jülich, Germany.
Eggeling
Lothar
L
Institute for Biotechnology, Research Centre Jülich, D-52425 Jülich, Germany.
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
D000995:Antitubercular Agents; D001426:Bacterial Proteins; D005971:Glutamates; D009171:Mycolic Acids; D011134:Polysaccharides; C030080:araban; D004977:Ethambutol; D010430:Pentosyltransferases; C101967:arabinosyltransferase