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Wiesenfeld Fellowship, Henry Wheeler Center for Emerging and Neglected Diseases","award":["CNCND-32305-47950"],"award-info":[{"award-number":["CNCND-32305-47950"]}]},{"name":"Graduate Research Fellowship Program, U.S. National Science Foundation","award":["DGE-1752814"],"award-info":[{"award-number":["DGE-1752814"]}]}],"content-domain":{"domain":["www.plosone.org"],"crossmark-restriction":false},"short-container-title":["PLoS One"],"abstract":"<jats:p>\n                    There is an urgent need to develop a more efficacious anti-tuberculosis vaccine as the current live-attenuated vaccine strain BCG fails to prevent pulmonary infection in adults. In this study, we leverage a synthetic biology approach to engineer BCG to produce more (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMBPP), an intermediate of bacterial\u2014but not host\u2014isoprenoid biosynthesis via the methylerythritol phosphate (MEP) pathway. HMBPP strongly activates and expands V\u03b39V\u03b42 T cells, which are unique to higher-order primates and protect against\n                    <jats:italic>Mycobacterium tuberculosis<\/jats:italic>\n                    infection. BCG has been engineered to produce specific ligands and antigens to some success; in contrast, our strategy exploits a self-nonself recognition mechanism in the host via HMBPP sensing, which has not been attempted before. To inform the design of our recombinant strains, we performed synteny analyses of &gt;63 mycobacterial species and found that isoprenoid biosynthetic genes are not operonic across all the 356 surveyed genomes, but some genes are frequently found in pairs. Thus, we generated synthetic loci with the goal of specifically overproducing HMBPP and tested the ability of these engineered strains to induce human V\u03b39V\u03b42 expansion in an\n                    <jats:italic>in vitro<\/jats:italic>\n                    stimulation assay. We found that BCG expressing a synthetic MEP locus significantly enhanced V\u03b39V\u03b42 T cell expansion over the wild-type vaccine strain, and overexpression of the HMBPP synthase GcpE alone potently induced V\u03b39V\u03b42 T cell expansion with no downregulation of other pathway genes. Together these engineered strains present two successful strategies to accumulate HMBPP and overcome feedback inhibition of the MEP pathway.\n                  <\/jats:p>","DOI":"10.1371\/journal.pone.0343925","type":"journal-article","created":{"date-parts":[[2026,4,6]],"date-time":"2026-04-06T17:26:36Z","timestamp":1775496396000},"page":"e0343925","update-policy":"https:\/\/doi.org\/10.1371\/journal.pone.corrections_policy","source":"Crossref","is-referenced-by-count":1,"title":["Leveraging a synthetic biology approach to enhance BCG-mediated expansion of V\u03b39V\u03b42 T cells"],"prefix":"10.1371","volume":"21","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-8345-9225","authenticated-orcid":true,"given":"Christine M.","family":"Qabar","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Allison W.","family":"Roberts","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4735-5285","authenticated-orcid":true,"given":"Lucas M.","family":"Waldburger","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Edward E. 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prospects","volume":"11","author":"L Zhuang","year":"2023","journal-title":"Vaccines"},{"key":"pone.0343925.ref002","doi-asserted-by":"crossref","first-page":"88","DOI":"10.1016\/j.smim.2018.07.001","article-title":"Genome wide approaches discover novel Mycobacterium tuberculosis antigens as correlates of infection, disease, immunity and targets for vaccination","volume":"39","author":"M Coppola","year":"2018","journal-title":"Semin Immunol"},{"issue":"5","key":"pone.0343925.ref003","doi-asserted-by":"crossref","first-page":"1274","DOI":"10.1128\/jb.178.5.1274-1282.1996","article-title":"Molecular analysis of genetic differences between Mycobacterium bovis BCG and virulent M. bovis","volume":"178","author":"GG Mahairas","year":"1996","journal-title":"J Bacteriol"},{"issue":"1","key":"pone.0343925.ref004","doi-asserted-by":"crossref","first-page":"117","DOI":"10.1086\/345862","article-title":"Deletion of RD1 from Mycobacterium tuberculosis mimics bacille Calmette-Gu\u00e9rin attenuation","volume":"187","author":"KN Lewis","year":"2003","journal-title":"J Infect Dis"},{"issue":"4","key":"pone.0343925.ref005","doi-asserted-by":"crossref","first-page":"470","DOI":"10.1093\/cid\/cit790","article-title":"Protection by BCG vaccine against tuberculosis: a systematic review of randomized controlled trials","volume":"58","author":"P Mangtani","year":"2014","journal-title":"Clin Infect Dis"},{"key":"pone.0343925.ref006","doi-asserted-by":"crossref","first-page":"316","DOI":"10.3389\/fimmu.2020.00316","article-title":"Vaccination against tuberculosis: revamping BCG by molecular genetics guided by immunology","volume":"11","author":"SHE Kaufmann","year":"2020","journal-title":"Front Immunol"},{"key":"pone.0343925.ref007","author":"WHO","year":"2019"},{"issue":"1","key":"pone.0343925.ref008","doi-asserted-by":"crossref","first-page":"878","DOI":"10.1038\/s41467-022-28509-z","article-title":"Re-engineered BCG overexpressing cyclic di-AMP augments trained immunity and exhibits improved efficacy against bladder cancer","volume":"13","author":"AK Singh","year":"2022","journal-title":"Nat Commun"},{"key":"pone.0343925.ref009","doi-asserted-by":"crossref","DOI":"10.7554\/eLife.89157","article-title":"A modified BCG with depletion of enzymes associated with peptidoglycan amidation induces enhanced protection against tuberculosis in mice","volume":"13","author":"MT Shaku","year":"2024","journal-title":"Elife"},{"issue":"1","key":"pone.0343925.ref010","doi-asserted-by":"crossref","first-page":"36","DOI":"10.1186\/s12951-021-01234-3","article-title":"Nanocages engineered from Bacillus Calmette-Guerin facilitate protective V\u03b32V\u03b42 T cell immunity against Mycobacterium tuberculosis infection","volume":"20","author":"J Pi","year":"2022","journal-title":"J Nanobiotechnology"},{"issue":"2","key":"pone.0343925.ref011","doi-asserted-by":"crossref","first-page":"468","DOI":"10.1038\/s41564-024-01895-4","article-title":"A BCG kill switch strain protects 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Karunakaran","year":"2014","journal-title":"Immunogenetics"},{"issue":"35","key":"pone.0343925.ref014","article-title":"Phosphoantigen-induced conformational change of butyrophilin 3A1 (BTN3A1) and its implication on V\u03b39V\u03b42 T cell activation","volume":"114","author":"S Gu","year":"2017","journal-title":"Proc Natl Acad Sci U S A"},{"issue":"6","key":"pone.0343925.ref015","doi-asserted-by":"crossref","first-page":"957","DOI":"10.1189\/jlb.6HI1213-632R","article-title":"HMBPP-deficient Listeria mutant immunization alters pulmonary\/systemic responses, effector functions, and memory polarization of V\u03b32V\u03b42 T cells","volume":"96","author":"JT Frencher","year":"2014","journal-title":"J Leukoc Biol"},{"issue":"19","key":"pone.0343925.ref016","doi-asserted-by":"crossref","DOI":"10.1172\/jci.insight.121899","article-title":"Mucosal-associated invariant and \u03b3\u03b4 T cell subsets respond to initial Mycobacterium tuberculosis infection","volume":"3","author":"CK Vorkas","year":"2018","journal-title":"JCI Insight"},{"key":"pone.0343925.ref017","doi-asserted-by":"crossref","first-page":"317","DOI":"10.1016\/S0014-5793(01)03191-X","article-title":"Identification of (E)\u20104\u2010hydroxy\u20103\u2010methyl\u2010but\u20102\u2010enyl pyrophosphate as a major activator for human \u03b3\u03b4 T cells in Escherichia coli","volume":"509","author":"M Hintz","year":"2001","journal-title":"FEBS Lett"},{"issue":"6","key":"pone.0343925.ref018","doi-asserted-by":"crossref","first-page":"3655","DOI":"10.4049\/jimmunol.166.6.3655","article-title":"Cutting edge: human gamma delta T cells are activated by intermediates of the 2-C-methyl-D-erythritol 4-phosphate pathway of isoprenoid biosynthesis","volume":"166","author":"B Altincicek","year":"2001","journal-title":"J Immunol"},{"issue":"7","key":"pone.0343925.ref019","doi-asserted-by":"crossref","first-page":"1257","DOI":"10.1016\/S0960-894X(03)00138-0","article-title":"Replacing the pyrophosphate group of HMB-PP by a diphosphonate function abrogates Its potential to activate human gammadelta T cells but does not lead to competitive antagonism","volume":"13","author":"A Reichenberg","year":"2003","journal-title":"Bioorg Med Chem Lett"},{"issue":"26","key":"pone.0343925.ref020","doi-asserted-by":"crossref","first-page":"14837","DOI":"10.1073\/pnas.201399298","article-title":"Studies on the nonmevalonate pathway to terpenes: the role of the GcpE (IspG) protein","volume":"98","author":"S Hecht","year":"2001","journal-title":"Proc Natl Acad Sci U S A"},{"issue":"2","key":"pone.0343925.ref021","doi-asserted-by":"crossref","first-page":"200","DOI":"10.1046\/j.1365-2567.2002.01414.x","article-title":"Accumulation of a potent gammadelta T-cell stimulator after deletion of the lytB gene in Escherichia coli","volume":"106","author":"M Eberl","year":"2002","journal-title":"Immunology"},{"issue":"20","key":"pone.0343925.ref022","doi-asserted-by":"crossref","first-page":"3780","DOI":"10.1021\/acs.accounts.1c00296","article-title":"Assembly-line catalysis in bifunctional terpene synthases","volume":"54","author":"JL Faylo","year":"2021","journal-title":"Acc Chem Res"},{"issue":"4","key":"pone.0343925.ref023","doi-asserted-by":"crossref","first-page":"293","DOI":"10.1007\/s11745-004-1233-3","article-title":"Isoprenoids: remarkable diversity of form and function","volume":"39","author":"SA Holstein","year":"2004","journal-title":"Lipids"},{"issue":"1","key":"pone.0343925.ref024","doi-asserted-by":"crossref","first-page":"19","DOI":"10.1042\/BJ20121899","article-title":"Metabolic plasticity for isoprenoid biosynthesis in bacteria","volume":"452","author":"J P\u00e9rez-Gil","year":"2013","journal-title":"Biochem J"},{"issue":"2","key":"pone.0343925.ref025","doi-asserted-by":"crossref","first-page":"249","DOI":"10.1039\/D1NP00047K","article-title":"Biosynthesis, evolution and ecology of microbial terpenoids","volume":"39","author":"M Avalos","year":"2022","journal-title":"Nat Prod Rep"},{"issue":"37","key":"pone.0343925.ref026","doi-asserted-by":"crossref","first-page":"23328","DOI":"10.1016\/S0021-9258(17)31657-5","article-title":"Recognition of the lipid intermediate for arabinogalactan\/arabinomannan biosynthesis and its relation to the mode of action of ethambutol on mycobacteria","volume":"269","author":"BA Wolucka","year":"1994","journal-title":"J Biol Chem"},{"issue":"22","key":"pone.0343925.ref027","doi-asserted-by":"crossref","first-page":"7564","DOI":"10.1128\/JB.186.22.7564-7570.2004","article-title":"Decaprenyl diphosphate synthesis in Mycobacterium tuberculosis","volume":"186","author":"D Kaur","year":"2004","journal-title":"J Bacteriol"},{"issue":"13","key":"pone.0343925.ref028","doi-asserted-by":"crossref","first-page":"7820","DOI":"10.1074\/jbc.271.13.7820","article-title":"Biosynthesis of the linkage region of the mycobacterial cell wall","volume":"271","author":"K Mikusov\u00e1","year":"1996","journal-title":"J Biol Chem"},{"issue":"1","key":"pone.0343925.ref029","doi-asserted-by":"crossref","first-page":"75","DOI":"10.1271\/bbb.100570","article-title":"Characterization of the Rv3378c gene product, a new diterpene synthase for producing tuberculosinol and (13R, S)-isotuberculosinol (nosyberkol), from the Mycobacterium tuberculosis H37Rv genome","volume":"75","author":"C Nakano","year":"2011","journal-title":"Biosci Biotechnol Biochem"},{"issue":"17","key":"pone.0343925.ref030","doi-asserted-by":"crossref","first-page":"3588","DOI":"10.1111\/j.1742-4658.2010.07767.x","article-title":"Diterpene production in Mycobacterium tuberculosis","volume":"277","author":"L Prach","year":"2010","journal-title":"FEBS J"},{"issue":"48","key":"pone.0343925.ref031","doi-asserted-by":"crossref","first-page":"17526","DOI":"10.1021\/ja9019287","article-title":"Edaxadiene: a new bioactive diterpene from Mycobacterium tuberculosis","volume":"131","author":"FM Mann","year":"2009","journal-title":"J Am Chem Soc"},{"issue":"7","key":"pone.0343925.ref032","doi-asserted-by":"crossref","first-page":"4798","DOI":"10.4049\/jimmunol.181.7.4798","article-title":"Definition of APC presentation of phosphoantigen (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate to Vgamma2Vdelta 2 TCR","volume":"181","author":"H Wei","year":"2008","journal-title":"J Immunol"},{"key":"pone.0343925.ref033","doi-asserted-by":"crossref","first-page":"5392","DOI":"10.1128\/IAI.01376-07","article-title":"Analysis of the isoprenoid biosynthesis pathways in Listeria monocytogenes reveals a role for the alternative 2-C-methyl-d-erythritol 4-phosphate pathway in murine infection","volume":"76","author":"M Begley","year":"2008","journal-title":"Infect Immun"},{"issue":"9","key":"pone.0343925.ref034","doi-asserted-by":"crossref","first-page":"2424","DOI":"10.1128\/JB.01402-09","article-title":"The nonmevalonate pathway of isoprenoid biosynthesis in Mycobacterium tuberculosis is essential and transcriptionally regulated by Dxs","volume":"192","author":"AC Brown","year":"2010","journal-title":"J Bacteriol"},{"issue":"6478","key":"pone.0343925.ref035","doi-asserted-by":"crossref","DOI":"10.1126\/science.aay5516","article-title":"Butyrophilin 2A1 is essential for phosphoantigen reactivity by \u03b3\u03b4 T cells","volume":"367","author":"M Rigau","year":"2020","journal-title":"Science"},{"issue":"5","key":"pone.0343925.ref036","doi-asserted-by":"crossref","first-page":"2390","DOI":"10.4049\/jimmunol.1401064","article-title":"Activation of human \u03b3\u03b4 T cells by cytosolic interactions of BTN3A1 with soluble phosphoantigens and the cytoskeletal adaptor periplakin","volume":"194","author":"DA Rhodes","year":"2015","journal-title":"J Immunol"},{"issue":"4","key":"pone.0343925.ref037","doi-asserted-by":"crossref","first-page":"490","DOI":"10.1016\/j.immuni.2014.03.003","article-title":"The intracellular B30.2 domain of butyrophilin 3A1 binds phosphoantigens to mediate activation of human V\u03b39V\u03b42 T cells","volume":"40","author":"A Sandstrom","year":"2014","journal-title":"Immunity"},{"key":"pone.0343925.ref038","doi-asserted-by":"crossref","first-page":"1973","DOI":"10.1016\/j.celrep.2016.04.081","article-title":"RhoB Mediates Phosphoantigen Recognition by V\u03b39V\u03b42 T Cell Receptor","volume":"15","author":"Z Sebestyen","year":"2016","journal-title":"Cell Rep"},{"issue":"11","key":"pone.0343925.ref039","doi-asserted-by":"crossref","first-page":"2269","DOI":"10.1182\/blood-2012-05-430470","article-title":"Key implication of CD277\/butyrophilin-3 (BTN3A) in cellular stress sensing by a major human \u03b3\u03b4 T-cell 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Infect Dis"},{"issue":"1","key":"pone.0343925.ref043","doi-asserted-by":"crossref","DOI":"10.1371\/journal.ppat.1003119","article-title":"Granzyme A produced by \u03b3(9)\u03b4(2) T cells induces human macrophages to inhibit growth of an intracellular pathogen","volume":"9","author":"CT Spencer","year":"2013","journal-title":"PLoS Pathog"},{"issue":"8","key":"pone.0343925.ref044","article-title":"Phosphoantigen\/IL2 expansion and differentiation of V\u03b32V\u03b42 T cells increase resistance to tuberculosis in nonhuman primates","volume":"9","author":"CY Chen","year":"2013","journal-title":"PLoS Pathog"},{"issue":"13","key":"pone.0343925.ref045","doi-asserted-by":"crossref","first-page":"6371","DOI":"10.1073\/pnas.1811380116","article-title":"Immunization of V\u03b32V\u03b42 T cells programs sustained effector memory responses that control tuberculosis in nonhuman primates","volume":"116","author":"L Shen","year":"2019","journal-title":"Proc Natl Acad Sci U S A"},{"key":"pone.0343925.ref046","doi-asserted-by":"crossref","first-page":"264","DOI":"10.1126\/science.1110267","article-title":"Professional antigen-presentation function by human \u03b3\u03b4 T cells","volume":"309","author":"M Brandes","year":"2005","journal-title":"Science"},{"key":"pone.0343925.ref047","doi-asserted-by":"crossref","first-page":"913","DOI":"10.3389\/fimmu.2019.00913","article-title":"IL-12 expands and differentiates human V\u03b32V\u03b42 T effector cells producing antimicrobial cytokines and inhibiting intracellular mycobacterial growth","volume":"10","author":"R Yang","year":"2019","journal-title":"Front Immunol"},{"issue":"6","key":"pone.0343925.ref048","doi-asserted-by":"crossref","first-page":"2249","DOI":"10.1084\/jem.178.6.2249","article-title":"An essential role for interferon gamma in resistance to Mycobacterium tuberculosis infection","volume":"178","author":"JL Flynn","year":"1993","journal-title":"J Exp Med"},{"issue":"1","key":"pone.0343925.ref049","doi-asserted-by":"crossref","first-page":"39","DOI":"10.1084\/jem.186.1.39","article-title":"Interleukin 12 (IL-12) is crucial to the development of protective immunity in mice intravenously infected with mycobacterium tuberculosis","volume":"186","author":"AM Cooper","year":"1997","journal-title":"J Exp Med"},{"issue":"5563","key":"pone.0343925.ref050","doi-asserted-by":"crossref","first-page":"2255","DOI":"10.1126\/science.1068819","article-title":"Adaptive immune response of Vgamma2Vdelta2+ T cells during mycobacterial infections","volume":"295","author":"Y Shen","year":"2002","journal-title":"Science"},{"issue":"12","key":"pone.0343925.ref051","doi-asserted-by":"crossref","first-page":"4753","DOI":"10.4049\/jimmunol.1602019","article-title":"Adoptive transfer of phosphoantigen-specific \u03b3\u03b4 T cell subset attenuates Mycobacterium tuberculosis infection in nonhuman primates","volume":"198","author":"A Qaqish","year":"2017","journal-title":"J Immunol"},{"issue":"2","key":"pone.0343925.ref052","doi-asserted-by":"crossref","first-page":"243","DOI":"10.1007\/s10295-019-02257-4","article-title":"A regulated synthetic operon facilitates stable overexpression of multigene terpenoid pathway in Bacillus subtilis","volume":"47","author":"II Abdallah","year":"2020","journal-title":"J Ind Microbiol Biotechnol"},{"key":"pone.0343925.ref053","doi-asserted-by":"crossref","first-page":"16274","DOI":"10.1038\/nmicrobiol.2016.274","article-title":"Programmable transcriptional repression in mycobacteria using an orthogonal CRISPR interference platform","volume":"2","author":"JM Rock","year":"2017","journal-title":"Nat Microbiol"},{"issue":"1","key":"pone.0343925.ref054","doi-asserted-by":"crossref","first-page":"60","DOI":"10.1038\/s41589-019-0400-9","article-title":"A computational framework to explore large-scale biosynthetic diversity","volume":"16","author":"JC Navarro-Mu\u00f1oz","year":"2020","journal-title":"Nat Chem Biol"},{"key":"pone.0343925.ref055","doi-asserted-by":"crossref","DOI":"10.1093\/nar\/gkab1112","article-title":"Database resources of the national center for biotechnology information","volume":"50","author":"EW Sayers","year":"2022","journal-title":"Nucleic Acids Res"},{"issue":"4","key":"pone.0343925.ref056","doi-asserted-by":"crossref","first-page":"366","DOI":"10.1038\/s41592-021-01101-x","article-title":"Sensitive protein alignments at tree-of-life scale using DIAMOND","volume":"18","author":"B Buchfink","year":"2021","journal-title":"Nat Methods"},{"issue":"6","key":"pone.0343925.ref057","doi-asserted-by":"crossref","first-page":"1101","DOI":"10.1038\/nprot.2008.73","article-title":"Analyzing real-time PCR data by the comparative C(T) method","volume":"3","author":"TD Schmittgen","year":"2008","journal-title":"Nat Protoc"},{"key":"pone.0343925.ref058","first-page":"579","article-title":"Metabolomics of Mycobacterium tuberculosis.","author":"KA Planck","year":"2021","journal-title":"Methods in Molecular Biology. Springer Protocols. Springer Nature"},{"issue":"1","key":"pone.0343925.ref059","doi-asserted-by":"crossref","first-page":"8","DOI":"10.1038\/s41540-024-00480-z","article-title":"Addressing genome scale design tradeoffs in Pseudomonas putida for bioconversion of an aromatic carbon source","volume":"11","author":"D Banerjee","year":"2025","journal-title":"NPJ Syst Biol Appl"},{"issue":"3","key":"pone.0343925.ref060","doi-asserted-by":"crossref","DOI":"10.1128\/spectrum.01158-21","article-title":"Genomic Degeneration and Reduction in the Fish Pathogen Mycobacterium shottsii","volume":"10","author":"DT Gauthier","year":"2022","journal-title":"Microbiol Spectr"},{"issue":"1","key":"pone.0343925.ref061","doi-asserted-by":"crossref","first-page":"12040","DOI":"10.1038\/s41598-018-30152-y","article-title":"Extensive genomic diversity among Mycobacterium marinum strains revealed by whole genome sequencing","volume":"8","author":"S Das","year":"2018","journal-title":"Sci Rep"},{"key":"pone.0343925.ref062","doi-asserted-by":"crossref","first-page":"258","DOI":"10.1186\/1471-2164-13-258","article-title":"On the origin of Mycobacterium ulcerans, the causative agent of Buruli ulcer","volume":"13","author":"KD Doig","year":"2012","journal-title":"BMC Genomics"},{"key":"pone.0343925.ref063","doi-asserted-by":"crossref","first-page":"78","DOI":"10.1186\/1471-2180-8-78","article-title":"Dxr is essential in Mycobacterium tuberculosis and fosmidomycin resistance is due to a lack of uptake","volume":"8","author":"AC Brown","year":"2008","journal-title":"BMC Microbiol"},{"issue":"8","key":"pone.0343925.ref064","doi-asserted-by":"crossref","DOI":"10.1371\/journal.pone.0135638","article-title":"LytB1 and LytB2 of Mycobacterium tuberculosis are not genetically redundant","volume":"10","author":"AC Brown","year":"2015","journal-title":"PLoS One"},{"issue":"4","key":"pone.0343925.ref065","doi-asserted-by":"crossref","DOI":"10.1128\/mSystems.00070-19","article-title":"Genomewide Assessment of Mycobacterium tuberculosis Conditionally Essential Metabolic Pathways","volume":"4","author":"Y Minato","year":"2019","journal-title":"mSystems"},{"issue":"1","key":"pone.0343925.ref066","doi-asserted-by":"crossref","first-page":"77","DOI":"10.1046\/j.1365-2958.2003.03425.x","article-title":"Genes required for mycobacterial growth defined by high density mutagenesis","volume":"48","author":"CM Sassetti","year":"2003","journal-title":"Mol Microbiol"},{"issue":"9","key":"pone.0343925.ref067","doi-asserted-by":"crossref","DOI":"10.1371\/journal.ppat.1002251","article-title":"High-resolution phenotypic profiling defines genes essential for mycobacterial growth and cholesterol catabolism","volume":"7","author":"JE Griffin","year":"2011","journal-title":"PLoS Pathog"},{"issue":"1","key":"pone.0343925.ref068","doi-asserted-by":"crossref","DOI":"10.1128\/mBio.02133-16","article-title":"Comprehensive Essentiality Analysis of the Mycobacterium tuberculosis Genome via Saturating Transposon Mutagenesis","volume":"8","author":"MA DeJesus","year":"2017","journal-title":"mBio"},{"key":"pone.0343925.ref069","doi-asserted-by":"crossref","first-page":"283","DOI":"10.1146\/annurev.genet.37.050503.084247","article-title":"Lateral gene transfer and the origins of prokaryotic groups","volume":"37","author":"Y Boucher","year":"2003","journal-title":"Annu Rev Genet"},{"issue":"29","key":"pone.0343925.ref070","doi-asserted-by":"crossref","DOI":"10.1073\/pnas.1105105108","article-title":"Evolution of the V, D, and J gene segments used in the primate gammadelta T-cell receptor reveals a dichotomy of conservation and diversity","volume":"108","author":"AR Kazen","year":"2011","journal-title":"Proc Natl Acad Sci U S A"},{"issue":"1","key":"pone.0343925.ref071","doi-asserted-by":"crossref","first-page":"5303","DOI":"10.1038\/s41467-024-49483-8","article-title":"The methylerythritol phosphate pathway as an oxidative stress sense and response system","volume":"15","author":"J Perez-Gil","year":"2024","journal-title":"Nat Commun"},{"issue":"23","key":"pone.0343925.ref072","doi-asserted-by":"crossref","first-page":"16926","DOI":"10.1074\/jbc.M113.464636","article-title":"Feedback inhibition of deoxy-D-xylulose-5-phosphate synthase regulates the methylerythritol 4-phosphate pathway","volume":"288","author":"A Banerjee","year":"2013","journal-title":"J Biol Chem"},{"key":"pone.0343925.ref073","doi-asserted-by":"crossref","first-page":"225","DOI":"10.1016\/bs.mie.2016.02.025","article-title":"Quantifying the Metabolites of the Methylerythritol 4-Phosphate (MEP) Pathway in Plants and Bacteria by Liquid Chromatography-Triple Quadrupole Mass Spectrometry","volume":"576","author":"D Gonz\u00e1lez-Cabanelas","year":"2016","journal-title":"Methods Enzymol"},{"key":"pone.0343925.ref074","first-page":"145","article-title":"Imaging the NADH:NAD+ Homeostasis for Understanding the Metabolic Response of Mycobacterium to Physiologically Relevant Stresses","volume":"6","author":"SA Bhat","year":"2016","journal-title":"Front Cell Infect Microbiol"},{"issue":"2","key":"pone.0343925.ref075","doi-asserted-by":"crossref","first-page":"444","DOI":"10.1042\/bj0870444","article-title":"Nicotin-amide-adenine nucleotides of Mycobacterium tuberculosis H37Rv","volume":"87","author":"KP Gopinathan","year":"1963","journal-title":"Biochem J"},{"issue":"33","key":"pone.0343925.ref076","doi-asserted-by":"crossref","first-page":"11945","DOI":"10.1073\/pnas.0711697105","article-title":"The protonmotive force is required for maintaining ATP homeostasis and viability of hypoxic, nonreplicating Mycobacterium tuberculosis","volume":"105","author":"SPS Rao","year":"2008","journal-title":"Proc Natl Acad Sci U S A"},{"issue":"28","key":"pone.0343925.ref077","doi-asserted-by":"crossref","first-page":"19329","DOI":"10.1074\/jbc.M800694200","article-title":"Biosynthesis and recycling of nicotinamide cofactors in mycobacterium tuberculosis. An essential role for NAD in nonreplicating bacilli","volume":"283","author":"HIM Boshoff","year":"2008","journal-title":"J Biol Chem"},{"issue":"4","key":"pone.0343925.ref078","article-title":"Metabolically distinct roles of NAD synthetase and NAD kinase define the essentiality of NAD and NADP in Mycobacterium tuberculosis","volume":"14","author":"R Sharma","year":"2023","journal-title":"mBio"},{"issue":"4","key":"pone.0343925.ref079","doi-asserted-by":"crossref","DOI":"10.1128\/mBio.01665-21","article-title":"Role of premycofactocin synthase in growth, microaerophilic adaptation, and metabolism of Mycobacterium tuberculosis","volume":"12","author":"G Krishnamoorthy","year":"2021","journal-title":"mBio"},{"key":"pone.0343925.ref080","doi-asserted-by":"crossref","first-page":"106974","DOI":"10.1016\/j.bioorg.2023.106974","article-title":"From bacteria to biomedicine: developing therapies exploiting NAD+ metabolism","volume":"142","author":"Y Chen","year":"2024","journal-title":"Bioorg Chem"},{"issue":"8","key":"pone.0343925.ref081","doi-asserted-by":"crossref","DOI":"10.1136\/jitc-2022-004880","article-title":"V\u03b42 T cells are associated with favorable clinical outcomes in patients with bladder cancer and their tumor reactivity can be boosted by BCG and zoledronate treatments","volume":"10","author":"S Nguyen","year":"2022","journal-title":"J Immunother Cancer"},{"issue":"44","key":"pone.0343925.ref082","doi-asserted-by":"crossref","first-page":"18374","DOI":"10.1021\/ja307315u","article-title":"Observation of thiamin-bound intermediates and microscopic rate constants for their interconversion on 1-deoxy-D-xylulose 5-phosphate synthase: 600-fold rate acceleration of pyruvate decarboxylation by D-glyceraldehyde-3-phosphate","volume":"134","author":"H Patel","year":"2012","journal-title":"J Am Chem Soc"}],"container-title":["PLOS One"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0343925","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,4,6]],"date-time":"2026-04-06T17:26:47Z","timestamp":1775496407000},"score":1,"resource":{"primary":{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pone.0343925"}},"subtitle":[],"editor":[{"given":"Atul","family":"Vashist","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2026,4,6]]},"references-count":82,"journal-issue":{"issue":"4","published-online":{"date-parts":[[2026,4,6]]}},"URL":"https:\/\/doi.org\/10.1371\/journal.pone.0343925","relation":{},"ISSN":["1932-6203"],"issn-type":[{"value":"1932-6203","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,4,6]]}}}