The mycobacterial DNA-binding protein 1 (MDP1) from Mycobacterium bovis BCG influences various growth characteristics.
BACKGROUND
Pathogenic mycobacteria such as M. tuberculosis, M. bovis or M. leprae are characterised by their extremely slow growth rate which plays an important role in mycobacterial virulence and eradication of the bacteria. Various limiting factors influence the generation time of mycobacteria, and the mycobacterial DNA-binding protein 1 (MDP1) has also been implicated in growth regulation. Our strategy to investigate the role of MDP1 in mycobacterial growth consisted in the generation and characterisation of a M. bovis BCG derivative expressing a MDP1-antisense gene.


RESULTS
The expression rate of the MDP1 protein in the recombinant M. bovis BCG containing the MDP1-antisense plasmid was reduced by about 50% compared to the reference strain M. bovis BCG containing the empty vector. In comparison to this reference strain, the recombinant M. bovis BCG grew faster in broth culture and reached higher cell masses in stationary phase. Likewise its intracellular growth in mouse and human macrophages was ameliorated. Bacterial clumping in broth culture was reduced by the antisense plasmid. The antisense plasmid increased the susceptibility of the bacteria towards Ampicillin. 2-D protein gels of bacteria maintained under oxygen-poor conditions demonstrated a reduction in the number and the intensity of many protein spots in the antisense strain compared to the reference strain.


CONCLUSIONS
The MDP1 protein has a major impact on various growth characteristics of M. bovis BCG. It plays an important role in virulence-related traits such as aggregate formation and intracellular multiplication. Its impact on the protein expression in a low-oxygen atmosphere indicates a role in the adaptation to the hypoxic conditions present in the granuloma.
Robert-Koch-Institut, Nordufer 20, 13353 Berlin, Germany. Lewina@rki.de
Astrid
A
Lewin
Daniela
D
Baus
Elisabeth
E
Kamal
Fabienne
F
Bon
Ralph
R
Kunisch
Sven
S
Maurischat
Michaela
M
Adonopoulou
Katharina
K
Eich
D000667:Ampicillin / Q000494:pharmacology; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D008264:Macrophages / Q000382:microbiology / Q000502:physiology; D051379:Mice; D009163:Mycobacterium bovis / Q000187:drug effects / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000235:genetics; D010100:Oxygen / Q000378:metabolism; D010957:Plasmids / Q000235:genetics
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C403010:MDP1 protein, Mycobacterium; D016376:Oligonucleotides, Antisense; D000667:Ampicillin; D010100:Oxygen