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Volume 160, Issue 7

Identification of a novel ATP-binding cassette transporter involved in long-chain fatty acid import and its role in triacylglycerol accumulation in RHA1 Free

Abstract

Members of the genus are specialists in the biosynthesis and accumulation of triacylglycerols (TAGs). As no transport protein related to TAG metabolism has yet been characterized in these bacteria, we used the available genomic information of RHA1 to perform a broad survey of genes coding for putative lipid transporter proteins in this oleaginous micro-organism. Among the seven genes encoding putative lipid transporters, (now called : lipid transporter protein) coding for an ATP-binding cassette protein was found clustered with others genes encoding enzymes catalysing the three putative acylation reactions of the Kennedy pathway for TAG synthesis. Overexpression of in the RHA1 strain led to an increase of approximately sixfold and threefold in biomass and TAG production, respectively, when cells were cultivated on palmitic acid and oleic acid. Moreover, overexpression of also promoted a significant increase in the uptake of a fluorescently labelled long-chain fatty acid (LCFA), as compared with the WT strain RHA1, and its further incorporation into the TAG fraction. Gluconate-grown cells showed increasing amounts of intracellular free fatty acids, but not of TAG, after overexpressing . Thus, for the first time to our knowledge, a transporter functionally related to TAG metabolism was identified in oleaginous rhodococci. Our results suggested that Ltp1 is an importer of LCFAs that plays a functional role in lipid homeostasis of RHA1.

  • Received:
  • Accepted:
  • Published Online:
Funding
This study was supported by the:
  • SCyT of the University of Patagonia San Juan Bosco
  • Agencia Comodoro Conocimiento (MCR)
  • Oil m&s Company
  • CONICET (Award PIP-CONICET no. 0764))
  • COFECyT (Award PFIP CHU-25)
  • ANPCyT (Award PICT2012 no. 2031)
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2014-07-01
2024-04-25
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References

  1. Alvarez H. M., Steinbüchel A. ( 2002). Triacylglycerols in prokaryotic microorganisms. Appl Microbiol Biotechnol 60:367–376 [View Article][PubMed]
     [Google Scholar]
  2. Alvarez H. M., Steinbüchel A. 2010; Physiology, bio-chemistry and molecular biology of triacylglycerol accumulation by Rhodococcus. Biology of RhodococcusMicrobiology Monographs vol. 16263–290 Alvarez H. M., Steinbüchel A. Heidelberg: Springer; [View Article]
     [Google Scholar]
  3. Alvarez H. M., Mayer F., Fabritius D., Steinbüchel A. ( 1996). Formation of intracytoplasmic lipid inclusions by Rhodococcus opacus strain PD630. Arch Microbiol 165:377–386 [View Article][PubMed]
     [Google Scholar]
  4. Alvarez A. F., Alvarez H. M., Kalscheuer R., Wältermann M., Steinbüchel A. ( 2008). Cloning and characterization of a gene involved in triacylglycerol biosynthesis and identification of additional homologous genes in the oleaginous bacterium Rhodococcus opacus PD630. Microbiology 154:2327–2335 [View Article][PubMed]
     [Google Scholar]
  5. Alvarez H. M., Silva R. A., Herrero M., Hernández A. M., Villalba M. S. ( 2013). Metabolism of triacylglycerols in Rhodococcus species: insights from physiology and molecular genetics. J Mol Biochem 2:69–78
     [Google Scholar]
  6. Aziz R. K., Bartels D., Best A. A., DeJongh M., Disz T., Edwards R. A., Formsma K., Gerdes S., Glass E. M. & other authors ( 2008). The RAST Server: rapid annotations using subsystems technology. BMC Genomics 9:75 [View Article][PubMed]
     [Google Scholar]
  7. Black P. N., DiRusso C. C. ( 2003). Transmembrane movement of exogenous long-chain fatty acids: proteins, enzymes, and vectorial esterification. Microbiol Mol Biol Rev 67:454–472 [View Article][PubMed]
     [Google Scholar]
  8. Borst P., Zelcer N., van Helvoort A. ( 2000). ABC transporters in lipid transport. Biochim Biophys Acta 1486:128–144 [View Article][PubMed]
     [Google Scholar]
  9. Brandl H., Gross R. A., Lenz R. W., Fuller R. C. ( 1988). Pseudomonas oleovorans as a source of poly(hydroxyalkanoates) for potential applications as biodegradable polyesters. Appl Environ Microbiol 54:1977–1982[PubMed]
     [Google Scholar]
  10. Chen Y., Ding Y., Yang L., Yu J., Liu G., Wang X., Zhang S., Yu D., Song L. & other authors ( 2013). Integrated omics study delineates the dynamics of lipid droplets in Rhodococcus opacus PD630. Nucleic Acids Res 42:1052–1064 [View Article][PubMed]
     [Google Scholar]
  11. Dassa E., Bouige P. ( 2001). The ABC of ABCs: a phylogenetic and functional classification of ABC systems in living organisms. Res Microbiol 152:211–229 [View Article][PubMed]
     [Google Scholar]
  12. Davidson A. L., Chen J. ( 2004). ATP-binding cassette transporters in bacteria. Annu Rev Biochem 73:241–268 [View Article][PubMed]
     [Google Scholar]
  13. Dean M., Hamon Y., Chimini G. ( 2001). The human ATP-binding cassette (ABC) transporter superfamily. J Lipid Res 42:1007–1017[PubMed]
     [Google Scholar]
  14. Ding Y., Yang L., Zhang S., Wang Y., Du Y., Pu J., Peng G., Chen Y., Zhang H. & other authors ( 2012). Identification of the major functional proteins of prokaryotic lipid droplets. J Lipid Res 53:399–411 [View Article][PubMed]
     [Google Scholar]
  15. Doshi R., Nguyen T., Chang G. ( 2013). Transporter-mediated biofuel secretion. Proc Natl Acad Sci U S A 110:7642–7647 [View Article][PubMed]
     [Google Scholar]
  16. Eckford P. D. W., Sharom F. J. ( 2008). Functional characterization of Escherichia coli MsbA: interaction with nucleotides and substrates. J Biol Chem 283:12840–12850 [View Article][PubMed]
     [Google Scholar]
  17. Hara H., Stewart G. R., Mohn W. W. ( 2010). Involvement of a novel ABC transporter and monoalkyl phthalate ester hydrolase in phthalate ester catabolism by Rhodococcus jostii RHA1. Appl Environ Microbiol 76:1516–1523 [View Article][PubMed]
     [Google Scholar]
  18. Hernández M. A., Arabolaza A., Rodríguez E., Gramajo H., Alvarez H. M. ( 2013). The atf2 gene is involved in triacylglycerol biosynthesis and accumulation in the oleaginous Rhodococcus opacus PD630. Appl Microbiol Biotechnol 97:2119–2130 [View Article][PubMed]
     [Google Scholar]
  19. Hettema E. H., van Roermund C. W. T., Distel B., van den Berg M., Vilela C., Rodrigues-Pousada C., Wanders R. J., Tabak H. F. ( 1996). The ABC transporter proteins Pat1 and Pat2 are required for import of long-chain fatty acids into peroxisomes of Saccharomyces cerevisiae. EMBO J 15:3813–3822[PubMed]
     [Google Scholar]
  20. Holder J. W., Ulrich J. C., DeBono A. C., Godfrey P. A., Desjardins C. A., Zucker J., Zeng Q., Leach A. L. B., Ghiviriga I. & other authors ( 2011). Comparative and functional genomics of Rhodococcus opacus PD630 for biofuels development. PLoS Genet 7:e1002219 [View Article][PubMed]
     [Google Scholar]
  21. Jorgensen J. H., Ferraro M. J. ( 2009). Antimicrobial susceptibility testing: a review of general principles and contemporary practices. Clin Infect Dis 49:1749–1755 [View Article][PubMed]
     [Google Scholar]
  22. Kalscheuer R., Arenskötter M., Steinbüchel A. ( 1999). Establishment of a gene transfer system for Rhodococcus opacus PD630 based on electroporation and its application for recombinant biosynthesis of poly(3-hydroxyalkanoic acids). Appl Microbiol Biotechnol 52:508–515 [View Article][PubMed]
     [Google Scholar]
  23. Kaul G., Pattan G. ( 2011). MsbA ATP-binding cassette (ABC) transporter of E. coli: structure and possible flippase mechanism. Indian J Biochem Biophys 48:7–13[PubMed]
     [Google Scholar]
  24. Kim S., Yamaoka Y., Ono H., Kim H., Shim D., Maeshima M., Martinoia E., Cahoon E. B., Nishida I., Lee Y. ( 2013). AtABCA9 transporter supplies fatty acids for lipid synthesis to the endoplasmic reticulum. Proc Natl Acad Sci U S A 110:773–778 [View Article][PubMed]
     [Google Scholar]
  25. LeBlanc J. C., Gonçalves E. R., Mohn W. W. ( 2008). Global response to desiccation stress in the soil actinomycete Rhodococcus jostii RHA1. Appl Environ Microbiol 74:2627–2636 [View Article][PubMed]
     [Google Scholar]
  26. MacEachran D. P., Sinskey A. J. ( 2013). The Rhodococcus opacus TadD protein mediates triacylgylycerol metabolism by regulating intracellular NAD(P)H pools. Microb Cell Fact 12:104 [View Article][PubMed]
     [Google Scholar]
  27. MacEachran D. P., Prophete M. E., Sinskey A. J. ( 2010). The Rhodococcus opacus PD630 heparin-binding hemagglutinin homolog TadA mediates lipid body formation. Appl Environ Microbiol 76:7217–7225 [View Article][PubMed]
     [Google Scholar]
  28. Maloy S. R., Ginsburgh C. L., Simons R. W., Nunn W. D. ( 1981). Transport of long and medium chain fatty acids by Escherichia coli K12. J Biol Chem 256:3735–3742[PubMed]
     [Google Scholar]
  29. Marmur J. ( 1961). A procedure for the isolation of deoxyribonucleic acid from microorganisms. J Mol Biol 3:208–218 [View Article]
     [Google Scholar]
  30. Marqués A. M., Pinazo A., Farfan M., Aranda F. J., Teruel J. A., Ortiz A., Manresa A., Espuny M. J. ( 2009). The physicochemical properties and chemical composition of trehalose lipids produced by Rhodococcus erythropolis 51T7. Chem Phys Lipids 158:110–117 [View Article][PubMed]
     [Google Scholar]
  31. McLeod M. P., Warren R. L., Hsiao W. W. L., Araki N., Myhre M., Fernandes C., Miyazawa D., Wong W., Lillquist A. L. & other authors ( 2006). The complete genome of Rhodococcus sp. RHA1 provides insights into a catabolic powerhouse. Proc Natl Acad Sci U S A 103:15582–15587 [View Article][PubMed]
     [Google Scholar]
  32. Nagao K., Kimura Y., Mastuo M., Ueda K. ( 2010). Lipid outward translocation by ABC proteins. FEBS Lett 584:2717–2723 [View Article][PubMed]
     [Google Scholar]
  33. Pohl A., Devaux P. F., Herrmann A. ( 2005). Function of prokaryotic and eukaryotic ABC proteins in lipid transport. Biochim Biophys Acta 1733:29–52 [View Article][PubMed]
     [Google Scholar]
  34. Sambrook J., Fritsch E. F., Maniatis T. ( 1989). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
     [Google Scholar]
  35. Schlegel H. G., Kaltwasser H., Gottschalk G. ( 1961). [A submersion method for culture of hydrogen-oxidizing bacteria: growth physiological studies]. Arch Mikrobiol 38:209–222 [View Article][PubMed]
     [Google Scholar]
  36. Schneider E., Hunke S. ( 1998). ATP-binding-cassette (ABC) transport systems: functional and structural aspects of the ATP-hydrolyzing subunits/domains. FEMS Microbiol Rev 22:1–20 [View Article][PubMed]
     [Google Scholar]
  37. Swain K., Casabon I., Eltis L. D., Mohn W. W. ( 2012). Two transporters essential for reassimilation of novel cholate metabolites by Rhodococcus jostii RHA1. J Bacteriol 194:6720–6727 [View Article][PubMed]
     [Google Scholar]
  38. van der Geize R., Hessels G. I., van Gerwen R., Vrijbloed J. W., van der Meijden P., Dijkhuizen L. ( 2000). Targeted disruption of the kstD gene encoding a 3-ketosteroid Δ1-dehydrogenase isoenzyme of Rhodococcus erythropolis strain SQ1. Appl Environ Microbiol 66:2029–2036 [View Article][PubMed]
     [Google Scholar]
  39. van Meer G., Halter D., Sprong H., Somerharju P., Egmond M. R. ( 2006). ABC lipid transporters: extruders, flippases, or flopless activators?. FEBS Lett 580:1171–1177 [View Article][PubMed]
     [Google Scholar]
  40. van Veen H. W., Venema K., Bolhuis H., Oussenko I., Kok J., Poolman B., Driessen A. J. M., Konings W. N. ( 1996). Multidrug resistance mediated by a bacterial homolog of the human multidrug transporter MDR1. Proc Natl Acad Sci U S A 93:10668–10672 [View Article][PubMed]
     [Google Scholar]
  41. Velamakanni S., Yao Y., Gutmann D. A. P., van Veen H. W. ( 2008). Multidrug transport by the ABC transporter Sav1866 from Staphylococcus aureus. Biochemistry 47:9300–9308 [View Article][PubMed]
     [Google Scholar]
  42. Villalba M. S., Hernández M. A., Silva R. A., Álvarez H. M. ( 2013). Genome sequences of triacylglycerol metabolism in Rhodococcus as a platform for comparative genomics. J Mol Biochem 2:94–105
     [Google Scholar]
  43. Wältermann M., Steinbüchel A. ( 2005). Neutral lipid bodies in prokaryotes: recent insights into structure, formation, and relationship to eukaryotic lipid depots. J Bacteriol 187:3607–3619 [View Article][PubMed]
     [Google Scholar]
  44. Wältermann M., Luftmann H., Baumeister D., Kalscheuer R., Steinbüchel A. ( 2000). Rhodococcus opacus strain PD630 as a new source of high-value single-cell oil? Isolation and characterization of triacylglycerols and other storage lipids. Microbiology 146:1143–1149[PubMed]
     [Google Scholar]
  45. Warhurst A. M., Fewson C. A. ( 1994). Biotransformations catalyzed by the genus Rhodococcus. Crit Rev Biotechnol 14:29–73 [View Article][PubMed]
     [Google Scholar]
  46. Zhou Z., White K. A., Polissi A., Georgopoulos C., Raetz C. R. H. ( 1998). Function of Escherichia coli MsbA, an essential ABC family transporter, in lipid A and phospholipid biosynthesis. J Biol Chem 273:12466–12475 [View Article][PubMed]
     [Google Scholar]
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Identification of a novel ATP-binding cassette transporter involved in long-chain fatty acid import and its role in triacylglycerol accumulation in Rhodococcus jostii RHA1
Microbiology 160, 1523 (2014); https://doi.org/10.1099/mic.0.078477-0
/content/journal/micro/10.1099/mic.0.078477-0
/content/journal/micro/10.1099/mic.0.078477-0
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