title: Small-molecule inhibitors of Staphylococcus aureus RnpA-mediated RNA turnover and tRNA processing.
abstract: New agents are urgently needed for the therapeutic treatment of Staphylococcus aureus infections. In that regard, S. aureus RNase RnpA may represent a promising novel dual-function antimicrobial target that participates in two essential cellular processes, RNA degradation and tRNA maturation. Accordingly, we previously used a high-throughput screen to identify small-molecule inhibitors of the RNA-degrading activity of the enzyme and showed that the RnpA inhibitor RNPA1000 is an attractive antimicrobial development candidate. In this study, we used a series of in vitro and cellular assays to characterize a second RnpA inhibitor, RNPA2000, which was identified in our initial screening campaign and is structurally distinct from RNPA1000. In doing so, it was found that S. aureus RnpA does indeed participate in 5'-precursor tRNA processing, as was previously hypothesized. Further, we show that RNPA2000 is a bactericidal agent that inhibits both RnpA-associated RNA degradation and tRNA maturation activities both in vitro and within S. aureus. The compound appears to display specificity for RnpA, as it did not significantly affect the in vitro activities of unrelated bacterial or eukaryotic ribonucleases and did not display measurable human cytotoxicity. Finally, we show that RNPA2000 exhibits antimicrobial activity and inhibits tRNA processing in efflux-deficient Gram-negative pathogens. Taken together, these data support the targeting of RnpA for antimicrobial development purposes, establish that small-molecule inhibitors of both of the functions of the enzyme can be identified, and lend evidence that RnpA inhibitors may have broad-spectrum antimicrobial activities.
authors.affiliation: Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska, USA Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry Rochester, New York, USA.
authors.forename: Tess M
authors.initials: TM
authors.lastname: Eidem
authors.affiliation: Moulder Center for Drug Discovery Research, Temple University, Philadelphia, Pennsylvania, USA.
authors.forename: Nicole
authors.initials: N
authors.lastname: Lounsbury
authors.affiliation: Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry Rochester, New York, USA.
authors.forename: John F
authors.initials: JF
authors.lastname: Emery
authors.affiliation: Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry Rochester, New York, USA.
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Bulger
authors.affiliation: Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry Rochester, New York, USA.
authors.forename: Andrew
authors.initials: A
authors.lastname: Smith
authors.affiliation: Moulder Center for Drug Discovery Research, Temple University, Philadelphia, Pennsylvania, USA.
authors.forename: Magid
authors.initials: M
authors.lastname: Abou-Gharbia
authors.affiliation: Moulder Center for Drug Discovery Research, Temple University, Philadelphia, Pennsylvania, USA wayne.childers@temple.edu paul_dunman@urmc.rochester.edu.
authors.forename: Wayne
authors.initials: W
authors.lastname: Childers
authors.affiliation: Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry Rochester, New York, USA wayne.childers@temple.edu paul_dunman@urmc.rochester.edu.
authors.forename: Paul M
authors.initials: PM
authors.lastname: Dunman
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D057166:High-Throughput Screening Assays; D006801:Humans; D006834:Hydrazines / Q000494:pharmacology; D008826:Microbial Sensitivity Tests; D012329:RNA, Bacterial / Q000187:drug effects*; D012343:RNA, Transfer / Q000187:drug effects*; D043262:Ribonuclease P / Q000037:antagonists & inhibitors*; D054852:Small Molecule Libraries; D013211:Staphylococcus aureus / Q000187:drug effects* / Q000378:metabolism*; D013890:Thiourea / Q000031:analogs & derivatives / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects
chemical_list: D000900:Anti-Bacterial Agents; D006834:Hydrazines; D012329:RNA, Bacterial; C000603507:RNPA2000; D054852:Small Molecule Libraries; D012343:RNA, Transfer; D043262:Ribonuclease P; D013890:Thiourea