Gain-of-Function Mutations in the Phospholipid Flippase MprF Confer Specific Daptomycin Resistance
ABSTRACT Daptomycin, a calcium-dependent lipopeptide antibiotic whose full mode of action is still not entirely understood, has become a standard-of-care agent for treating methicillin-resistant Staphylococcus aureus (MRSA) infections. Daptomycin-resistant (DAP-R) S. aureus mutants emerge during the...
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| Format: | Article |
| Language: | English |
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American Society for Microbiology
2018-12-01
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| Series: | mBio |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/mBio.01659-18 |
| _version_ | 1818856709687345152 |
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| author | Christoph M. Ernst Christoph J. Slavetinsky Sebastian Kuhn Janna N. Hauser Mulugeta Nega Nagendra N. Mishra Cordula Gekeler Arnold S. Bayer Andreas Peschel |
| author_facet | Christoph M. Ernst Christoph J. Slavetinsky Sebastian Kuhn Janna N. Hauser Mulugeta Nega Nagendra N. Mishra Cordula Gekeler Arnold S. Bayer Andreas Peschel |
| author_sort | Christoph M. Ernst |
| collection | DOAJ |
| description | ABSTRACT Daptomycin, a calcium-dependent lipopeptide antibiotic whose full mode of action is still not entirely understood, has become a standard-of-care agent for treating methicillin-resistant Staphylococcus aureus (MRSA) infections. Daptomycin-resistant (DAP-R) S. aureus mutants emerge during therapy, featuring isolates which in most cases possess point mutations in the mprF gene. MprF is a bifunctional bacterial resistance protein that synthesizes the positively charged lipid lysyl-phosphatidylglycerol (LysPG) and translocates it subsequently from the inner membrane leaflet to the outer membrane leaflet. This process leads to increased positive S. aureus surface charge and reduces susceptibility to cationic antimicrobial peptides and cationic antibiotics. We characterized the most commonly reported MprF mutations in DAP-R S. aureus strains in a defined genetic background and found that only certain mutations, including the frequently reported T345A single nucleotide polymorphism (SNP), can reproducibly cause daptomycin resistance. Surprisingly, T345A did not alter LysPG synthesis, LysPG translocation, or the S. aureus cell surface charge. MprF-mediated DAP-R relied on a functional flippase domain and was restricted to daptomycin and a related cyclic lipopeptide antibiotic, friulimicin B, suggesting that the mutations modulate specific interactions with these two antibiotics. Notably, the T345A mutation led to weakened intramolecular domain interactions of MprF, suggesting that daptomycin and friulimicin resistance-conferring mutations may alter the substrate range of the MprF flippase to directly translocate these lipopeptide antibiotics or other membrane components with crucial roles in the activity of these antimicrobials. Our study points to a new mechanism used by S. aureus to resist calcium-dependent lipopeptide antibiotics and increases our understanding of the bacterial phospholipid flippase MprF. IMPORTANCE Ever since daptomycin was introduced to the clinic, daptomycin-resistant isolates have been reported. In most cases, the resistant isolates harbor point mutations in MprF, which produces and flips the positively charged phospholipid LysPG. This has led to the assumption that the resistance mechanism relies on the overproduction of LysPG, given that increased LysPG production may lead to increased electrostatic repulsion of positively charged antimicrobial compounds, including daptomycin. Here we show that the resistance mechanism is highly specific and relies on a different process that involves a functional MprF flippase, suggesting that the resistance-conferring mutations may enable the flippase to accommodate daptomycin or an unknown component that is crucial for its activity. Our report provides a new perspective on the mechanism of resistance to a major antibiotic. |
| first_indexed | 2024-12-19T08:28:49Z |
| format | Article |
| id | doaj.art-91185988fd2b4f0eadcc6a36be94023e |
| institution | Directory Open Access Journal |
| issn | 2150-7511 |
| language | English |
| last_indexed | 2024-12-19T08:28:49Z |
| publishDate | 2018-12-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | mBio |
| spelling | doaj.art-91185988fd2b4f0eadcc6a36be94023e2022-12-21T20:29:15ZengAmerican Society for MicrobiologymBio2150-75112018-12-019610.1128/mBio.01659-18Gain-of-Function Mutations in the Phospholipid Flippase MprF Confer Specific Daptomycin ResistanceChristoph M. Ernst0Christoph J. Slavetinsky1Sebastian Kuhn2Janna N. Hauser3Mulugeta Nega4Nagendra N. Mishra5Cordula Gekeler6Arnold S. Bayer7Andreas Peschel8Interfaculty Institute of Microbiology and Infection Medicine, Infection Biology, University of Tübingen, Tübingen, GermanyInterfaculty Institute of Microbiology and Infection Medicine, Infection Biology, University of Tübingen, Tübingen, GermanyInterfaculty Institute of Microbiology and Infection Medicine, Infection Biology, University of Tübingen, Tübingen, GermanyInterfaculty Institute of Microbiology and Infection Medicine, Infection Biology, University of Tübingen, Tübingen, GermanyInterfaculty Institute of Microbiology and Infection Medicine, Infection Biology, University of Tübingen, Tübingen, GermanyLos Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, Torrance, California, USAInterfaculty Institute of Microbiology and Infection Medicine, Infection Biology, University of Tübingen, Tübingen, GermanyLos Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, Torrance, California, USAInterfaculty Institute of Microbiology and Infection Medicine, Infection Biology, University of Tübingen, Tübingen, GermanyABSTRACT Daptomycin, a calcium-dependent lipopeptide antibiotic whose full mode of action is still not entirely understood, has become a standard-of-care agent for treating methicillin-resistant Staphylococcus aureus (MRSA) infections. Daptomycin-resistant (DAP-R) S. aureus mutants emerge during therapy, featuring isolates which in most cases possess point mutations in the mprF gene. MprF is a bifunctional bacterial resistance protein that synthesizes the positively charged lipid lysyl-phosphatidylglycerol (LysPG) and translocates it subsequently from the inner membrane leaflet to the outer membrane leaflet. This process leads to increased positive S. aureus surface charge and reduces susceptibility to cationic antimicrobial peptides and cationic antibiotics. We characterized the most commonly reported MprF mutations in DAP-R S. aureus strains in a defined genetic background and found that only certain mutations, including the frequently reported T345A single nucleotide polymorphism (SNP), can reproducibly cause daptomycin resistance. Surprisingly, T345A did not alter LysPG synthesis, LysPG translocation, or the S. aureus cell surface charge. MprF-mediated DAP-R relied on a functional flippase domain and was restricted to daptomycin and a related cyclic lipopeptide antibiotic, friulimicin B, suggesting that the mutations modulate specific interactions with these two antibiotics. Notably, the T345A mutation led to weakened intramolecular domain interactions of MprF, suggesting that daptomycin and friulimicin resistance-conferring mutations may alter the substrate range of the MprF flippase to directly translocate these lipopeptide antibiotics or other membrane components with crucial roles in the activity of these antimicrobials. Our study points to a new mechanism used by S. aureus to resist calcium-dependent lipopeptide antibiotics and increases our understanding of the bacterial phospholipid flippase MprF. IMPORTANCE Ever since daptomycin was introduced to the clinic, daptomycin-resistant isolates have been reported. In most cases, the resistant isolates harbor point mutations in MprF, which produces and flips the positively charged phospholipid LysPG. This has led to the assumption that the resistance mechanism relies on the overproduction of LysPG, given that increased LysPG production may lead to increased electrostatic repulsion of positively charged antimicrobial compounds, including daptomycin. Here we show that the resistance mechanism is highly specific and relies on a different process that involves a functional MprF flippase, suggesting that the resistance-conferring mutations may enable the flippase to accommodate daptomycin or an unknown component that is crucial for its activity. Our report provides a new perspective on the mechanism of resistance to a major antibiotic.https://journals.asm.org/doi/10.1128/mBio.01659-18daptomycinMRSAMprFStaphylococcus aureusantibiotic resistanceflippase |
| spellingShingle | Christoph M. Ernst Christoph J. Slavetinsky Sebastian Kuhn Janna N. Hauser Mulugeta Nega Nagendra N. Mishra Cordula Gekeler Arnold S. Bayer Andreas Peschel Gain-of-Function Mutations in the Phospholipid Flippase MprF Confer Specific Daptomycin Resistance mBio daptomycin MRSA MprF Staphylococcus aureus antibiotic resistance flippase |
| title | Gain-of-Function Mutations in the Phospholipid Flippase MprF Confer Specific Daptomycin Resistance |
| title_full | Gain-of-Function Mutations in the Phospholipid Flippase MprF Confer Specific Daptomycin Resistance |
| title_fullStr | Gain-of-Function Mutations in the Phospholipid Flippase MprF Confer Specific Daptomycin Resistance |
| title_full_unstemmed | Gain-of-Function Mutations in the Phospholipid Flippase MprF Confer Specific Daptomycin Resistance |
| title_short | Gain-of-Function Mutations in the Phospholipid Flippase MprF Confer Specific Daptomycin Resistance |
| title_sort | gain of function mutations in the phospholipid flippase mprf confer specific daptomycin resistance |
| topic | daptomycin MRSA MprF Staphylococcus aureus antibiotic resistance flippase |
| url | https://journals.asm.org/doi/10.1128/mBio.01659-18 |
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