title: The nonmevalonate pathway of isoprenoid biosynthesis in Mycobacterium tuberculosis is essential and transcriptionally regulated by Dxs.
abstract: Mycobacterium tuberculosis synthesizes isoprenoids via the nonmevalonate or DOXP pathway. Previous work demonstrated that three enzymes in the pathway (Dxr, IspD, and IspF) are all required for growth in vitro. We demonstrate the essentiality of the key genes dxs1 and gcpE, confirming that the pathway is of central importance and that the second homolog of the synthase (dxs2) cannot compensate for the loss of dxs1. We looked at the effect of overexpression of Dxr, Dxs1, Dxs2, and GcpE on viability and on growth in M. tuberculosis. Overexpression of dxs1 or dxs2 was inhibitory to growth, whereas overexpression of dxr or gcpE was not. Toxicity is likely to be, at least partially, due to depletion of pyruvate from the cells. Overexpression of dxs1 or gcpE resulted in increased flux through the pathway, as measured by accumulation of the metabolite 4-hydroxy-3-methyl-but-2-enyl pyrophosphate. We identified the functional translational start site and promoter region for dxr and demonstrated that it is expressed as part of a polycistronic mRNA with gcpE and two other genes. Increased expression of this operon was seen in cells overexpressing Dxs1, indicating that transcriptional control is effected by the first enzyme of the pathway via an unknown regulator.
authors.affiliation: Queen Mary University of London, Barts and the London School of Medicine and Dentistry, London E12AD, United Kingdom.
authors.forename: Amanda C
authors.initials: AC
authors.lastname: Brown
authors.forename: Matthias
authors.initials: M
authors.lastname: Eberl
authors.forename: Dean C
authors.initials: DC
authors.lastname: Crick
authors.forename: Hassan
authors.initials: H
authors.lastname: Jomaa
authors.forename: Tanya
authors.initials: T
authors.lastname: Parish
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D011756:Diphosphates / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics / Q000502:physiology; D050296:Microbial Viability / Q000235:genetics; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D013729:Terpenes / Q000378:metabolism*
chemical_list: C443995:4-hydroxy-3-methylbut-2-enyl pyrophosphate; D001426:Bacterial Proteins; D011756:Diphosphates; D013729:Terpenes