Efflux-mediated bis-indole resistance in Staphylococcus aureus reveals differential substrate specificities for MepA and MepR.
The bis-indoles are a novel class of compounds with potent antibacterial activity against a broad spectrum of Gram-positive and Gram-negative pathogens. The mechanism of action of these compounds has not been clearly defined. To study the mechanism of action of bis-indoles, selections for mutants of Staphylococcus aureus NCTC 8325 with reduced susceptibility to several chemically related bis-indoles were carried out using serial passages in subinhibitory compound concentrations. Resistant mutants were only obtained for one of the four bis-indoles tested (MBX-1090), and these appeared at concentrations up to 16X MIC within 10-12 passages. MBX-1090 resistance mutations produced a truncated open reading frame of mepR (SAOUHSC_00314), a gene encoding a MarR-like repressor. MepR regulates expression of mepA (SAOUHSC_00315), which encodes a member of the Multidrug and Toxic Compound Extrusion (MATE) family of efflux pumps. MBX-1090 resistance was reverted when mepR (wild type) was provided in trans. Microarray experiments and RT-PCR experiments confirmed that over-expression of mepA is required for resistance. Interestingly, MBX-1090 resistant mutants and strains overexpressing mepA from an expression vector did not exhibit cross-resistance to closely related bis-indole compounds. MBX-1090 did not induce expression of mepA, suggesting that this compound does not directly interact with MepR. Conversely, the bis-indoles that were not substrates of MepA strongly induced mepA expression. The results of this study suggest that MepA and MepR exhibit remarkably distinct substrate specificity for closely related bis-indoles.
Opperman
Timothy J
TJ
Microbiotix, Inc., One Innovation Dr., Worcester, MA 01605-4332, United States.
Williams
John D
JD
Microbiotix, Inc., One Innovation Dr., Worcester, MA 01605-4332, United States.
Houseweart
Chad
C
Microbiotix, Inc., One Innovation Dr., Worcester, MA 01605-4332, United States.
Panchal
Rekha G
RG
United States Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Fort Detrick, Frederick, MA 21702-5011, United States.
Bavari
Sina
S
United States Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Fort Detrick, Frederick, MA 21702-5011, United States.
Peet
Norton P
NP
Microbiotix, Inc., One Innovation Dr., Worcester, MA 01605-4332, United States.
Moir
Donald T
DT
Microbiotix, Inc., One Innovation Dr., Worcester, MA 01605-4332, United States.
Bowlin
Terry L
TL
Microbiotix, Inc., One Innovation Dr., Worcester, MA 01605-4332, United States.
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D024881:Drug Resistance, Bacterial / Q000235:genetics; D007211:Indoles / Q000737:chemistry / Q000494:pharmacology*; D008826:Microbial Sensitivity Tests; D009154:Mutation; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013211:Staphylococcus aureus / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D001426:Bacterial Proteins; D007211:Indoles; D012097:Repressor Proteins