%0 Journal Article %A Rhodes, Katherine A. %A Somprasong, Nawarat %A Podnecky, Nicole L. %A Mima, Takehiko %A Chirakul, Sunisa %A Schweizer, Herbert P. %T Molecular determinants of Burkholderia pseudomallei BpeEF-OprC efflux pump expression %D 2018 %J Microbiology, %V 164 %N 9 %P 1156-1167 %@ 1465-2080 %R https://doi.org/10.1099/mic.0.000691 %K efflux %K Burkholderia pseudomallei %K antibiotic resistance %K melioidosis %K regulation %I Microbiology Society, %X Burkholderia pseudomallei, the cause of melioidosis, is intrinsically resistant to many antibiotics. Acquired multidrug resistance, including resistance to doxycycline and co-trimoxazole used for melioidosis eradication phase therapy, is mainly attributed to constitutive expression of the BpeEF-OprC efflux pump. Constitutive expression of this pump is caused by mutations affecting two highly similar LysR-type transcriptional regulators (LTTR), BpeT and BpeS, but their interaction with the regulatory region governing BpeEF-OprC expression has not yet been studied. The bpeE-bpeF-oprC genes are distally located in the llpE-bpeE-bpeF-oprC operon. The llpE gene encodes a putative lipase/esterase of unknown function. We show that in a bpeT mutant llpE is constitutively co-transcribed with bpeE-bpeF-oprC. As expected from previous studies with B. cenocepacia, deletion of llpE does not affect antibiotic efflux. Using transcriptional bpeE′-lacZ fusions, we demonstrate that the 188 bp bpeT-llpE intergenic region located between bpeT and the llpE-bpeE-bpeF-oprC operon contains regulatory elements needed for control of bpeT and llpE-bpeE-bpeF-oprC operon expression. By native polyacrylamide gel electrophoresis and electrophoretic mobility shift assays with purified recombinant BpeT and BpeS proteins, we show BpeT and BpeS form oligomers that share a 14 bp binding site overlapping the essential region required for llpE-bpeE-bpeF-oprC expression. The binding site contains the conserved T-N11-A LTTR box motif involved in binding of LysR proteins, which in concert with two other possible LTTR boxes may mediate BpeT and BpeS regulation of BpeEF-OprC expression. These studies form the basis for further investigation of BpeEF-OprC expression and regulation at the molecular level by yet unknown external stimuli. %U https://www.microbiologyresearch.org/content/journal/micro/10.1099/mic.0.000691