Inactivation of farR Causes High Rhodomyrtone Resistance and Increased Pathogenicity in Staphylococcus aureus

Abstract

Rhodomyrtone (Rom) is an acylphloroglucinol antibiotic originally isolated from leaves of Rhodomyrtus tomentosa . Rom targets the bacterial membrane and is active against a wide range of Gram-positive bacteria but the exact mode of action remains obscure. Here we isolated and characterized a spontaneous Rom-resistant mutant from the model strain Staphylococcus aureus HG001 (Rom R ) to learn more about the resistance mechanism. We showed that Rom-resistance is based on a single point mutation in the coding region of farR [regulator of f atty a cid (FA) r esistance] that causes an amino acid change from Cys to Arg at position 116 in FarR, that affects FarR activity. Comparative transcriptome analysis revealed that mutated farR affects transcription of many genes in distinct pathways. FarR represses for example the expression of its own gene ( farR ), its flanking gene farE (effector of FA resistance), and other global regulators such as agr and sarA . All these genes were consequently upregulated in the Rom R clone. Particularly the upregulation of agr and sarA leads to increased expression of virulence genes rendering the Rom R clone more cytotoxic and more pathogenic in a mouse infection model. The Rom-resistance is largely due to the de-repression of farE . FarE is described as an efflux pump for linoleic and arachidonic acids. We observed an increased release of lipids in the Rom R clone compared to its parental strain HG001. If farE is deleted in the Rom R clone, or, if native farR is expressed in the Rom R strain, the corresponding strains become hypersensitive to Rom. Overall, we show here that the high Rom-resistance is mediated by overexpression of farE in the Rom R clone, that FarR is an important regulator, and that the point mutation in farR (Rom R clone) makes the clone hyper-virulent.

Staphylococcus aureus Releases Proinflammatory Membrane Vesicles To Resist Antimicrobial Fatty Acids

“…FarE mediates AFA efflux under the control of its transcriptional regulator FarR (20). Constitutive activation of FarE confers increased resistance against AFAs and the membrane-targeting antibiotic rhodomyrtone (20,50). Remarkably, high FarE levels also lead to an increased release of PSMs (50).…”

“…Constitutive activation of FarE confers increased resistance against AFAs and the membrane-targeting antibiotic rhodomyrtone ( 20 , 50 ). Remarkably, high FarE levels also lead to an increased release of PSMs ( 50 ). Since PSMs are known to promote MV release ( 28 , 51 ), we surmise an indirect contribution of FarE to this process.…”

Efflux Pump Mediated Antimicrobial Resistance by Staphylococci in Health-Related Environments: Challenges and the Quest for Inhibition

“…In addition, highly conserved residues Y59 and W62 in TMS 3 were found to be functionally important as substitutions resulted in a significant reduction in the ability of QacC to confer resistance to benzalkonium and other compounds [ 145 ]. The qacC gene [ 130 , 150 ] can be carried on small rolling-circle replicating plasmids [ 151 , 152 ] and large low-copy-number multiresistance plasmids; its promoter is known to differ between these plasmids systems [ 130 ]. QacC expression is not modulated by a transcriptional regulator [ 141 ] and currently there is no high-resolution structure available for QacC protein.…”

Molecular Basis of Rhodomyrtone Resistance in Staphylococcus aureus

“…In our previous study, we were able to isolate Rom-resistant mutants by subculturing S. aureus HG001 in medium supplemented with Rom ( 12 ) and to attribute Rom resistance to a single point mutation in the coding region of farR (regulator of fatty acid resistance). FarR belongs to the TetR family of regulators (TFRs), which are widely associated with antibiotic resistance and the regulation of genes encoding small-molecule exporters, but they are also involved in controlling many other aspects of prokaryotic physiology ( 13 ).…”