Fitness cost and interference of Arm/Rmt aminoglycoside resistance methyltransferases with the RsmF housekeeping methyltransferase
May 1st, 2012
Investigation article published in Antimicrobial Agents and Chemotherapy
Arm/Rmt methyltransferases have emerged recently in pathogenic bacteria as enzymes that confer high-level resistance to 4,6 disubstituted aminoglycosides through methylation of the G1405 residue in the 16S rRNA. In prokaryotes, nucleotide methylations are the most common type of rRNA modification, and are introduced post- transcriptionally by a variety of site-specific housekeeping enzymes to optimize ribosomal function. Here we show that while the aminoglycoside resistance methyltransferase RmtC methylates G1405, it impedes methylation of the housekeeping methyltranferase RsmF at position C1407, a nucleotide that, like G1405, forms part of the aminoglycoside binding pocket of the 16S rRNA. To understand the origin and consequences of this phenomenon, we constructed a series of in frame knock-out and knock-in mutants in Escherichia coli, corresponding to the genotypes rsmF(+)/rmtC(-), rsmF(-)/rmtC(-), rsmF(+)/rmtC(+) and rsmF(-)/rmtC(+). When analyzed for the antimicrobial resistance pattern, the rsmF(-) bacteria presented a decreased susceptibility to aminoglycosides, including 4,6- and 4,5-deoxystreptamine aminoglycosides, showing that the housekeeping methylation at C1407 is involved in intrinsic aminoglycoside resistance in E. coli. Competition experiments between the isogenic E. coli strains showed that, contrary to expectation, acquisition of rmtC does not entail fitness cost for the bacterium. Finally, MALDI mass spectrometry allowed us to determine that RmtC methylates the G1405 residue not only in presence, but also in absence of aminoglycoside antibiotics. Thus, the coupling between housekeeping and acquired methyltransferases subverts the methylation architecture of the 16S rRNA, but elicits Arm/Rmt methyltransferases to be selected and retained, posing an important threat to the usefulness of aminoglycosides world-wide
Gutierrez B., Escudero JA., San Millan A., Hidalgo L., Carrilero L., Ovejero CM., Santos-Lopez A., Thomas-Lopez D. and Gonzalez-Zorn B..