1887

Abstract

Lipid metabolism is critical to Mycobacterium tuberculosis survival and infection. Unlike Escherichia coli, which has a single FadR, the M. tuberculosis genome encodes five proteins of the FadR sub-family. While the role of E. coli FadR as a regulator of fatty acid metabolism is well known, the definitive functions of M. tuberculosis FadR proteins are still under investigation. An interesting question about the M. tuberculosis FadRs remains open: which one of these proteins is the functional homologue of E. coli FadR? To address this, we have applied two different approaches. The first one was the bioinformatics approach and the second one was the classical molecular genetic approach involving complementation studies. Surprisingly, the results of these two approaches did not agree. Among the five M. tuberculosis FadRs, Rv0494 shared the highest sequence similarity with FadR and Rv0586 was the second best match. However, only Rv0586, but not Rv0494, could complement E. coli ∆fadR, indicating that Rv0586 is the M. tuberculosis functional homologue of FadR . Further studies showed that both regulators, Rv0494 and Rv0586, show similar responsiveness to LCFA, and have conserved critical residues for DNA binding. However, analysis of the operator site indicated that the inter-palindromic distance required for DNA binding differs for the two regulators. The differences in the binding site selection helped in the success of Rv0586 binding to fadB upstream over Rv0494 and may have played a critical role in complementing E. coli ∆fadR. Further, for the first time, we report the lipid-responsive nature of Rv0586.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.000686
2018-07-11
2019-12-15
Loading full text...

Full text loading...

/deliver/fulltext/micro/164/9/1133.html?itemId=/content/journal/micro/10.1099/mic.0.000686&mimeType=html&fmt=ahah

References

  1. Vindal V, Ranjan S, Ranjan A. In silico analysis and characterization of GntR family of regulators from Mycobacterium tuberculosis. Tuberculosis 2007;87:242–247 [CrossRef][PubMed]
    [Google Scholar]
  2. Dirusso CC, Heimert TL, Metzger AK. Characterization of FadR, a global transcriptional regulator of fatty acid metabolism in Escherichia coli. Interaction with the fadB promoter is prevented by long chain fatty acyl coenzyme A. J Biol Chem 1992;267:8685–8691[PubMed]
    [Google Scholar]
  3. Dirusso CC, Metzger AK, Heimert TL. Regulation of transcription of genes required for fatty acid transport and unsaturated fatty acid biosynthesis in Escherichia coli by FadR. Mol Microbiol 1993;7:311–322 [CrossRef][PubMed]
    [Google Scholar]
  4. Gui L, Sunnarborg A, Laporte DC. Regulated expression of a repressor protein: FadR activates iclR. J Bacteriol 1996;178:4704–4709 [CrossRef][PubMed]
    [Google Scholar]
  5. Farewell A, Diez AA, Dirusso CC, Nyström T. Role of the Escherichia coli FadR regulator in stasis survival and growth phase-dependent expression of the uspA, fad, and fab genes. J Bacteriol 1996;178:6443–6450 [CrossRef][PubMed]
    [Google Scholar]
  6. Campbell JW, Cronan JE. Escherichia coli FadR positively regulates transcription of the fabB fatty acid biosynthetic gene. J Bacteriol 2001;183:5982–5990 [CrossRef][PubMed]
    [Google Scholar]
  7. Hoskisson PA, Rigali S, Fowler K, Findlay KC, Buttner MJ. DevA, a GntR-like transcriptional regulator required for development in Streptomyces coelicolor. J Bacteriol 2006;188:5014–5023 [CrossRef][PubMed]
    [Google Scholar]
  8. Georgi T, Engels V, Wendisch VF. Regulation of L-lactate utilization by the FadR-type regulator LldR of Corynebacterium glutamicum. J Bacteriol 2008;190:963–971 [CrossRef][PubMed]
    [Google Scholar]
  9. Feng Y, Cronan JE. The Vibrio cholerae fatty acid regulatory protein, FadR, represses transcription of plsB, the gene encoding the first enzyme of membrane phospholipid biosynthesis. Mol Microbiol 2011;81:1020–1033 [CrossRef][PubMed]
    [Google Scholar]
  10. Brown RN, Gulig PA. Regulation of fatty acid metabolism by FadR is essential for Vibrio vulnificus to cause infection of mice. J Bacteriol 2008;190:7633–7644 [CrossRef][PubMed]
    [Google Scholar]
  11. Cronan JE. In vivo evidence that acyl coenzyme A regulates DNA binding by the Escherichia coli FadR global transcription factor. J Bacteriol 1997;179:1819–1823 [CrossRef][PubMed]
    [Google Scholar]
  12. Henry MF, Cronan JE. A new mechanism of transcriptional regulation: release of an activator triggered by small molecule binding. Cell 1992;70:671–679 [CrossRef][PubMed]
    [Google Scholar]
  13. Dirusso CC, Tsvetnitsky V, Højrup P, Knudsen J. Fatty acyl-CoA binding domain of the transcription factor FadR. Characterization by deletion, affinity labeling, and isothermal titration calorimetry. J Biol Chem 1998;273:33652–33659[PubMed]
    [Google Scholar]
  14. Iram SH, Cronan JE. Unexpected functional diversity among FadR fatty acid transcriptional regulatory proteins. J Biol Chem 2005;280:32148–32156 [CrossRef][PubMed]
    [Google Scholar]
  15. van Aalten DM, Dirusso CC, Knudsen J, Wierenga RK. Crystal structure of FadR, a fatty acid-responsive transcription factor with a novel acyl coenzyme A-binding fold. Embo J 2000;19:5167–5177 [CrossRef][PubMed]
    [Google Scholar]
  16. van Aalten DM, Dirusso CC, Knudsen J. The structural basis of acyl coenzyme A-dependent regulation of the transcription factor FadR. Embo J 2001;20:2041–2050 [CrossRef][PubMed]
    [Google Scholar]
  17. Sartain MJ, Dick DL, Rithner CD, Crick DC, Belisle JT. Lipidomic analyses of Mycobacterium tuberculosis based on accurate mass measurements and the novel "Mtb LipidDB". J Lipid Res 2011;52:861–872 [CrossRef][PubMed]
    [Google Scholar]
  18. Gatfield J, Pieters J. Essential role for cholesterol in entry of mycobacteria into macrophages. Science 2000;288:1647–1651 [CrossRef][PubMed]
    [Google Scholar]
  19. Lee W, Vanderven BC, Fahey RJ, Russell DG. Intracellular Mycobacterium tuberculosis exploits host-derived fatty acids to limit metabolic stress. J Biol Chem 2013;288:6788–6800 [CrossRef][PubMed]
    [Google Scholar]
  20. Anand S, Singh V, Singh AK, Mittal M, Datt M et al. Equilibrium binding and kinetic characterization of putative tetracycline repressor family transcription regulator Fad35R from Mycobacterium tuberculosis. Febs J 2012;279:3214–3228 [CrossRef][PubMed]
    [Google Scholar]
  21. Salzman V, Mondino S, Sala C, Cole ST, Gago G et al. Transcriptional regulation of lipid homeostasis in mycobacteria. Mol Microbiol 2010;78:64–77 [CrossRef][PubMed]
    [Google Scholar]
  22. Vindal V, Suma K, Ranjan A. GntR family of regulators in Mycobacterium smegmatis: a sequence and structure based characterization. BMC Genomics 2007;8:289 [CrossRef][PubMed]
    [Google Scholar]
  23. Biswas RK, Dutta D, Tripathi A, Feng Y, Banerjee M et al. Identification and characterization of Rv0494: a fatty acid-responsive protein of the GntR/FadR family from Mycobacterium tuberculosis. Microbiology 2013;159:913–923 [CrossRef][PubMed]
    [Google Scholar]
  24. Yousuf S, Angara R, Vindal V, Ranjan A. Rv0494 is a starvation-inducible, auto-regulatory FadR-like regulator from Mycobacterium tuberculosis. Microbiology 2015;161:463–476 [CrossRef][PubMed]
    [Google Scholar]
  25. Casali N, White AM, Riley LW. Regulation of the Mycobacterium tuberculosis mce1 operon. J Bacteriol 2006;188:441–449 [CrossRef][PubMed]
    [Google Scholar]
  26. Cantrell SA, Leavell MD, Marjanovic O, Iavarone AT, Leary JA et al. Free mycolic acid accumulation in the cell wall of the mce1 operon mutant strain of Mycobacterium tuberculosis. J Microbiol 2013;51:619–626 [CrossRef][PubMed]
    [Google Scholar]
  27. Cheigh C, Senaratne R, Uchida Y, Casali N, Kendall LV et al. Posttreatment reactivation of tuberculosis in mice caused by Mycobacterium tuberculosis disrupted in mce1R. 2010;202752–759
  28. Uchida Y, Casali N, White A, Morici L, Kendall LV et al. Accelerated immunopathological response of mice infected with Mycobacterium tuberculosis disrupted in the mce1 operon negative transcriptional regulator. Cell Microbiol 2007;9:1275–1283 [CrossRef][PubMed]
    [Google Scholar]
  29. Vindal V, Ashwantha Kumar E, Ranjan A. Identification of operator sites within the upstream region of the putative mce2R gene from mycobacteria. FEBS Lett 2008;582:1117–1122 [CrossRef][PubMed]
    [Google Scholar]
  30. Forrellad MA, Bianco MV, Blanco FC, Nuñez J, Klepp LI et al. Study of the in vivo role of Mce2R, the transcriptional regulator of mce2 operon in Mycobacterium tuberculosis. BMC Microbiol 2013;13:200 [CrossRef][PubMed]
    [Google Scholar]
  31. Santangelo ML, Blanco F, Campos E, Soria M, Bianco MV et al. Mce2R from Mycobacterium tuberculosis represses the expression of the mce2 operon. Tuberculosis 2009;89:22–28 [CrossRef][PubMed]
    [Google Scholar]
  32. Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y et al. Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol Syst Biol 2006;2:2006.0008 [CrossRef][PubMed]
    [Google Scholar]
  33. Sambrook J, Fritsch EF, Maniatis T. Molecular Cloning: A Laboratory Manual New York: Cold Spring Harbor Laboratory Press; 1989
    [Google Scholar]
  34. Pathania A, Sardesai AA. Distinct paths for basic amino acid export in Escherichia coli: YbjE (LysO) mediates export of L-lysine. J Bacteriol 2015;197:2036–2047 [CrossRef][PubMed]
    [Google Scholar]
  35. Edgar RC. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res 2004;32:1792–1797 [CrossRef][PubMed]
    [Google Scholar]
  36. Nguyen LT, Schmidt HA, Von Haeseler A, Minh BQ. IQ-TREE: a fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Mol Biol Evol 2015;32:268–274 [CrossRef][PubMed]
    [Google Scholar]
  37. Minh BQ, Nguyen MA, Von Haeseler A. Ultrafast approximation for phylogenetic bootstrap. Mol Biol Evol 2013;30:1188–1195 [CrossRef][PubMed]
    [Google Scholar]
  38. He Z, Zhang H, Gao S, Lercher MJ, Chen WH et al. Evolview v2: an online visualization and management tool for customized and annotated phylogenetic trees. Nucleic Acids Res 2016;44:W236–W241 [CrossRef][PubMed]
    [Google Scholar]
  39. Raman N, Black PN, Dirusso CC. Characterization of the fatty acid-responsive transcription factor FadR. Biochemical and genetic analyses of the native conformation and functional domains. J Biol Chem 1997;272:30645–30650[PubMed]
    [Google Scholar]
  40. Rigali S, Derouaux A, Giannotta F, Dusart J. Subdivision of the helix-turn-helix GntR family of bacterial regulators in the FadR, HutC, MocR, and YtrA subfamilies. J Biol Chem 2002;277:12507–12515 [CrossRef][PubMed]
    [Google Scholar]
  41. Nunn WD, Giffin K, Clark D, Cronan JE. Role for fadR in unsaturated fatty acid biosynthesis in Escherichia coli. J Bacteriol 1983;154:554–560[PubMed]
    [Google Scholar]
  42. Cole ST, Brosch R, Parkhill J, Garnier T, Churcher C et al. Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature 1998;393:537–544 [CrossRef][PubMed]
    [Google Scholar]
  43. Minch KJ, Rustad TR, Peterson EJ, Winkler J, Reiss DJ et al. The DNA-binding network of Mycobacterium tuberculosis. Nat Commun 2015;6:5829 [CrossRef][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.000686
Loading
/content/journal/micro/10.1099/mic.0.000686
Loading

Data & Media loading...

Supplements

Supplementary File 1

PDF
This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error