Mutations in dnaA and a cryptic interaction site increase drug resistance in Mycobacterium tuberculosis |
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Authors: | Nathan D Hicks Samantha R Giffen Peter H Culviner Michael C Chao Charles L Dulberger Qingyun Liu Sydney Stanley Jessica Brown Jaimie Sixsmith Ian D Wolf Sarah M Fortune |
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Institution: | 1. Department of Immunology and Infectious Diseases, Harvard T.H. Chan School of Public Health, Boston, Massachusetts, United States of America;2. Ragon Institute of MGH, MIT, and Harvard, Cambridge, Massachusetts, United States of America;3. Broad Institute of MIT and Harvard, Cambridge, Massachusetts, United States of America;National Institutes of Health, UNITED STATES |
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Abstract: | Genomic dissection of antibiotic resistance in bacterial pathogens has largely focused on genetic changes conferring growth above a single critical concentration of drug. However, reduced susceptibility to antibiotics—even below this breakpoint—is associated with poor treatment outcomes in the clinic, including in tuberculosis. Clinical strains of Mycobacterium tuberculosis exhibit extensive quantitative variation in antibiotic susceptibility but the genetic basis behind this spectrum of drug susceptibility remains ill-defined. Through a genome wide association study, we show that non-synonymous mutations in dnaA, which encodes an essential and highly conserved regulator of DNA replication, are associated with drug resistance in clinical M. tuberculosis strains. We demonstrate that these dnaA mutations specifically enhance M. tuberculosis survival during isoniazid treatment via reduced expression of katG, the activator of isoniazid. To identify DnaA interactors relevant to this phenotype, we perform the first genome-wide biochemical mapping of DnaA binding sites in mycobacteria which reveals a DnaA interaction site that is the target of recurrent mutation in clinical strains. Reconstructing clinically prevalent mutations in this DnaA interaction site reproduces the phenotypes of dnaA mutants, suggesting that clinical strains of M. tuberculosis have evolved mutations in a previously uncharacterized DnaA pathway that quantitatively increases resistance to the key first-line antibiotic isoniazid. Discovering genetic mechanisms that reduce drug susceptibility and support the evolution of high-level drug resistance will guide development of biomarkers capable of prospectively identifying patients at risk of treatment failure in the clinic. |
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