title: Elucidating the regulon of multidrug resistance regulator RarA in Klebsiella pneumoniae.
abstract: RarA is an AraC-type regulator in Klebsiella pneumoniae, which, when overexpressed, confers a low-level multidrug-resistant (MDR) phenotype linked to the upregulation of both the acrAB and oqxAB efflux genes. Increased rarA expression has also been shown to be integral in the development of tigecycline resistance in the absence of ramA in K. pneumoniae. Given its phenotypic role in MDR, microarray analyses were performed to determine the RarA regulon. Transcriptome analysis was undertaken using strains Ecl8ΔrarA/pACrarA-2 (rarA-expressing construct) and Ecl8ΔrarA/pACYC184 (vector-only control) using bespoke microarray slides consisting of probes derived from the genomic sequences of K. pneumoniae MGH 78578 (NC_009648.1) and Kp342 (NC_011283.1). Our results show that rarA overexpression resulted in the differential expression of 66 genes (42 upregulated and 24 downregulated). Under the COG (clusters of orthologous groups) functional classification, the majority of affected genes belonged to the category of cell envelope biogenesis and posttranslational modification, along with genes encoding the previously uncharacterized transport proteins (e.g., KPN_03141, sdaCB, and leuE) and the porin OmpF. However, genes associated with energy production and conversion and amino acid transport/metabolism (e.g., nuoA, narJ, and proWX) were found to be downregulated. Biolog phenotype analyses demonstrated that rarA overexpression confers enhanced growth of the overexpresser in the presence of several antibiotic classes (i.e., beta-lactams and fluoroquinolones), the antifungal/antiprotozoal compound clioquinol, disinfectants (8-hydroxyquinoline), protein synthesis inhibitors (i.e., minocycline and puromycin), membrane biogenesis agents (polymyxin B and amitriptyline), DNA synthesis (furaltadone), and the cytokinesis inhibitor (sanguinarine). Both our transcriptome and phenotypic microarray data support and extend the role of RarA in the MDR phenotype of K. pneumoniae.
authors: Centre for Infection and Immunity, Queen's University Belfast, Belfast, United Kingdom.
authors: Shyamasree
authors: S
authors: De Majumdar
authors: Mark
authors: M
authors: Veleba
authors: Sarah
authors: S
authors: Finn
authors: Séamus
authors: S
authors: Fanning
authors: Thamarai
authors: T
authors: Schneiders
mesh_terms: D047151:Amaryllidaceae Alkaloids / Q000494:pharmacology; D000639:Amitriptyline / Q000494:pharmacology; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D024841:Fluoroquinolones / Q000494:pharmacology; D007711:Klebsiella pneumoniae / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D008826:Microbial Sensitivity Tests; D009581:Nitrofurans / Q000494:pharmacology; D023303:Oxazolidinones / Q000494:pharmacology; D011112:Polymyxin B / Q000494:pharmacology; D011691:Puromycin / Q000494:pharmacology; D018085:Regulon / Q000235:genetics* / Q000502:physiology
chemical_list: D047151:Amaryllidaceae Alkaloids; D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D024841:Fluoroquinolones; D009581:Nitrofurans; D023303:Oxazolidinones; D000639:Amitriptyline; D011691:Puromycin; C007795:furaltadon; D011112:Polymyxin B; C419423:sanguinine