title: Aminoglycoside resistance 16S rRNA methyltransferases block endogenous methylation, affect translation efficiency and fitness of the host.
abstract: In Gram-negative bacteria, acquired 16S rRNA methyltransferases ArmA and NpmA confer high-level resistance to all clinically useful aminoglycosides by modifying, respectively, G1405 and A1408 in the A-site. These enzymes must coexist with several endogenous methyltransferases that are essential for fine-tuning of the decoding center, such as RsmH and RsmI in Escherichia coli, which methylate C1402 and RsmF C1407. The resistance methyltransferases have a contrasting distribution--ArmA has spread worldwide, whereas a single clinical isolate producing NpmA has been reported. The rate of dissemination of resistance depends on the fitness cost associated with its expression. We have compared ArmA and NpmA in isogenic Escherichia coli harboring the corresponding structural genes and their inactive point mutants cloned under the control of their native constitutive promoter in the stable plasmid pGB2. Growth rate determination and competition experiments showed that ArmA had a fitness cost due to methylation of G1405, whereas NpmA conferred only a slight disadvantage to the host due to production of the enzyme. MALDI MS indicated that ArmA impeded one of the methylations at C1402 by RsmI, and not at C1407 as previously proposed, whereas NpmA blocked the activity of RsmF at C1407. A dual luciferase assay showed that methylation at G1405 and A1408 and lack of methylation at C1407 affect translation accuracy. These results indicate that resistance methyltransferases impair endogenous methylation with different consequences on cell fitness.
authors.forename: Virginia S
authors.initials: VS
authors.lastname: Lioy
authors.forename: Sylvie
authors.initials: S
authors.lastname: Goussard
authors.forename: Vincent
authors.initials: V
authors.lastname: Guerineau
authors.forename: Eun-Jeong
authors.initials: EJ
authors.lastname: Yoon
authors.forename: Patrice
authors.initials: P
authors.lastname: Courvalin
authors.forename: Marc
authors.initials: M
authors.lastname: Galimand
authors.forename: Catherine
authors.initials: C
authors.lastname: Grillot-Courvalin
mesh_terms: D000617:Aminoglycosides / Q000494:pharmacology*; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D056084:Genetic Fitness; D008745:Methylation / Q000187:drug effects*; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D014176:Protein Biosynthesis / Q000187:drug effects*; D012333:RNA, Messenger / Q000235:genetics; D012336:RNA, Ribosomal, 16S / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D029968:Escherichia coli Proteins; D012333:RNA, Messenger; D012336:RNA, Ribosomal, 16S; D008780:Methyltransferases
keywords: 16S rRNA methyltransferase; aminoglycoside resistance; fitness; translation