Ioerger, Thomas R.O’Malley, TheresaLiao, ReilingGuinn, Kristine M.Hickey, Mark J.Mohaideen, NilofarMurphy, Kenan C.Boshoff, Helena I. M.Mizrahi, ValerieRubin, EricSassetti, Christopher M.Barry, Clifton E.Sherman, David R.Parish, TanyaSacchettini, James C.2014-03-102013Ioerger, T. R., T. O’Malley, R. Liao, K. M. Guinn, M. J. Hickey, N. Mohaideen, K. C. Murphy, et al. 2013. “Identification of New Drug Targets and Resistance Mechanisms in Mycobacterium tuberculosis.” PLoS ONE 8 (9): e75245. doi:10.1371/journal.pone.0075245. http://dx.doi.org/10.1371/journal.pone.0075245.1932-6203http://nrs.harvard.edu/urn-3:HUL.InstRepos:11878840Identification of new drug targets is vital for the advancement of drug discovery against Mycobacterium tuberculosis, especially given the increase of resistance worldwide to first- and second-line drugs. Because traditional target-based screening has largely proven unsuccessful for antibiotic discovery, we have developed a scalable platform for target identification in M. tuberculosis that is based on whole-cell screening, coupled with whole-genome sequencing of resistant mutants and recombineering to confirm. The method yields targets paired with whole-cell active compounds, which can serve as novel scaffolds for drug development, molecular tools for validation, and/or as ligands for co-crystallization. It may also reveal other information about mechanisms of action, such as activation or efflux. Using this method, we identified resistance-linked genes for eight compounds with anti-tubercular activity. Four of the genes have previously been shown to be essential: AspS, aspartyl-tRNA synthetase, Pks13, a polyketide synthase involved in mycolic acid biosynthesis, MmpL3, a membrane transporter, and EccB3, a component of the ESX-3 type VII secretion system. AspS and Pks13 represent novel targets in protein translation and cell-wall biosynthesis. Both MmpL3 and EccB3 are involved in membrane transport. Pks13, AspS, and EccB3 represent novel candidates not targeted by existing TB drugs, and the availability of whole-cell active inhibitors greatly increases their potential for drug discovery.en-USJournal Article2014-03-1010.1371/journal.pone.0075245