The transcription factor CwiR is important for envelope integrity in Burkholderia thailandensis

ABSTRACT: The genus Burkholderia contains more than 100 species, including opportunistic human pathogens, facultative intracellular pathogens, and phytopathogens. Burkholderia thailandensis is a surrogate for studying host-pathogen interactions, virulence mechanisms, and antimicrobial resistance within the Burkholderia genus on account of its limited virulence in humans. The Gram-negative Burkholderia species are highly resistant to a plethora of antimicrobial agents, in part due to efflux pumps, which play a central role in resistance mechanisms by exporting antimicrobial substances. An annotated operon comprising BTH_I2558-I261 was characterized. BTH_I2558 encodes a member of the multiple antibiotic resistance (MarR) family of transcription factors, which was named CwiR based on inferred roles in maintenance of cell wall integrity, and the other three genes (BTH_I2559-61) encode a tripartite efflux pump including an outer membrane protein, a periplasmic protein, and an inner membrane transporter protein. BTH_I2558-I261 was confirmed to be an operon using cDNA synthesis and PCR techniques. Real-time quantitative PCR and mRNA sequencing were carried out for gene expression analysis, and proteomic analysis was conducted using quantitative mass spectrometry. Using a ΔcwiR strain in which BTH_I2558 was disrupted, we found that CwiR represses the expression of the three adjacent efflux pump genes. Upregulation of these genes would be predicted to correlate with more antibiotics or other toxic compounds being pumped out of the bacterial cell. Unexpectedly, the ΔcwiR strain exhibited increased sensitivity to several tested antibiotics, suggesting that CwiR not only regulates the adjacent efflux pump genes but also controls other genes. RNA sequencing of the ΔcwiR strain revealed that CwiR serves as a master regulator that regulates 273 genes, including several involved in cell wall integrity. Consistent with this observation, elevated levels of proteins associated with structural maintenance of the cell wall were detected in the ΔcwiR proteome. Therefore, our data suggest that CwiR regulates antibiotic sensitivity by modifying cell wall integrity. This knowledge will aid in the development of new therapeutic interventions for multidrug resistant pathogenic Burkholderia species.

INSTRUMENT(S):

ORGANISM(S): Burkholderia Thailandensis E264 Bacteria

SUBMITTER: Ahmed Al-Tohamy  

LAB HEAD: Anne Grove

PROVIDER: PXD069908 | Pride | 2026-10-08

REPOSITORIES: Pride