1887

Abstract

Transhydrogenases catalyse interconversion of the redox cofactors NADH and NADPH, thereby conveying metabolic flexibility to balance catabolic NADPH formation with anabolic or stress-based consumption of NADPH. is one of the very few microbes that possesses two isoforms: the membrane-bound, proton-translocating transhydrogenase PntAB and the cytosolic, energy-independent transhydrogenase UdhA. Despite their physiological relevance, we have only fragmented information on their regulation and the signals coordinating their counteracting activities. Here we investigated PntAB and UdhA regulation by studying transcriptional responses to environmental and genetic perturbations. By testing and GFP reporter constructs in the background of WT and 62 transcription factor mutants during growth on different carbon sources, we show distinct transcriptional regulation of the two transhydrogenase promoters. Surprisingly, transhydrogenase regulation was independent of the actual catabolic overproduction or underproduction of NADPH but responded to nutrient levels and growth rate in a fashion that matches the cellular need for the redox cofactors NADPH and/or NADH. Specifically, the identified transcription factors Lrp, ArgP and Crp link transhydrogenase expression to particular amino acids and intracellular concentrations of cAMP. The overall identified set of regulators establishes a primarily biosynthetic role for PntAB and link UdhA to respiration.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.000346
2016-09-01
2024-03-28
Loading full text...

Full text loading...

/deliver/fulltext/micro/162/9/1672.html?itemId=/content/journal/micro/10.1099/mic.0.000346&mimeType=html&fmt=ahah

References

  1. Alton N. K., Vapnek D. 1979; Nucleotide sequence analysis of the chloramphenicol resistance transposon Tn9 . Nature 282:864–869 [View Article][PubMed]
    [Google Scholar]
  2. Baba T., Ara T., Hasegawa M., Takai Y., Okumura Y., Baba M., Datsenko K. A., Tomita M., Wanner B. L., Mori H. 2006; Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol Syst Biol 2:1 [View Article]
    [Google Scholar]
  3. Bakker B. M., Overkamp K. M., van Maris A. J., Kötter P., Luttik M. A., van Dijken J. P., Pronk J. T. 2001; Stoichiometry and compartmentation of NADH metabolism in Saccharomyces cerevisiae . FEMS Microbiol Rev 25:15–37 [View Article][PubMed]
    [Google Scholar]
  4. Bettenbrock K., Sauter T., Jahreis K., Kremling A., Lengeler J. W., Gilles E.-D. 2007; Correlation between growth rates, EIIACrr phosphorylation, and intracellular cyclic AMP levels in Escherichia coli K-12. J Bacteriol 189:6891–6900 [View Article][PubMed]
    [Google Scholar]
  5. Blank L. M., Lehmbeck F., Sauer U. 2005; Metabolic-flux and network analysis in fourteen hemiascomycetous yeasts. FEMS Yeast Res 5:545–558 [View Article][PubMed]
    [Google Scholar]
  6. Boonstra B., French C. E., Wainwright I., Bruce N. C. 1999; The udhA gene of Escherichia coli encodes a soluble pyridine nucleotide transhydrogenase. J Bacteriol 181:1030–1034[PubMed]
    [Google Scholar]
  7. Bouvier J., Stragier P., Morales V., Rémy E., Gutierrez C. 2008; Lysine represses transcription of the Escherichia coli dapB gene by preventing its activation by the ArgP activator. J Bacteriol 190:5224–5229 [View Article][PubMed]
    [Google Scholar]
  8. Calvo J. M., Matthews R. G. 1994; The leucine-responsive regulatory protein, a global regulator of metabolism in Escherichia coli . Microbiol Mol Biol Rev 58:466–490
    [Google Scholar]
  9. Chechik G., Oh E., Rando O., Weissman J., Regev A., Koller D. 2008; Activity motifs reveal principles of timing in transcriptional control of the yeast metabolic network. Nat Biotechnol 26:1251–1259 [View Article]
    [Google Scholar]
  10. Cho B.-K., Barrett C. L., Knight E. M., Park Y. S., Palsson B. Ø. 2008; Genome-scale reconstruction of the Lrp regulatory network in Escherichia coli . Proc Natl Acad Sci USA 105:19462–19467 [View Article][PubMed]
    [Google Scholar]
  11. Chou H. H., Marx C. J., Sauer U. 2015; Transhydrogenase promotes the robustness and evolvability of E. coli deficient in NADPH production. PLoS Genet 11:e1005007 [View Article][PubMed]
    [Google Scholar]
  12. Cui Y., Midkiff M. A., Wang Q., Calvo J. M. 1996; The leucine-responsive regulatory protein (Lrp) from Escherichia coli . J Biol Chem 271:6611–6617 [CrossRef]
    [Google Scholar]
  13. David M. C., Tip W. L., Shirley G., Philip D. B. 1986; Nucleotide sequence of the pntA and pntB genes encoding the pyridine nucleotide transhydrogenase of Escherichia coli . Eur J Biochem 158:647–653 [CrossRef]
    [Google Scholar]
  14. Doan T., Servant P., Tojo S., Yamaguchi H., Lerondel G., Yoshida K., Fujita Y., Aymerich S. 2003; The Bacillus subtilis ywkA gene encodes a malic enzyme and its transcription is activated by the YufL/YufM two-component system in response to malate. Microbiology 149:2331–2343 [View Article][PubMed]
    [Google Scholar]
  15. Driscoll B. T., Finan T. M. 1997; Properties of NAD+- and NADP+-dependent malic enzymes of Rhizobium (Sinorhizobium) meliloti and differential expression of their genes in nitrogen-fixing bacteroids. Microbiology 143:489–498 [View Article][PubMed]
    [Google Scholar]
  16. Edgar R., Domrachev M., Lash A. E. 2002; Gene Expression Omnibus: NCBI gene expression and hybridization array data repository. Nucleic Acids Res 30:207–210 [View Article][PubMed]
    [Google Scholar]
  17. Faith J. J., Driscoll M. E., Fusaro V. A., Cosgrove E. J., Hayete B., Juhn F. S., Schneider S. J., Gardner T. S. 2008; Many Microbe Microarrays Database: uniformly normalized Affymetrix compendia with structured experimental metadata. Nucleic Acids Res 36:D866–D870 [View Article][PubMed]
    [Google Scholar]
  18. Federowicz S., Kim D., Ebrahim A., Lerman J., Nagarajan H., Cho B.-K., Zengler K., Palsson B. 2014; Determining the control circuitry of redox metabolism at the genome-scale. PLoS Genet 10:e1004264 [View Article][PubMed]
    [Google Scholar]
  19. Fischer E., Sauer U. 2003; A novel metabolic cycle catalyzes glucose oxidation and anaplerosis in hungry Escherichia coli . J Biol Chem 278:46446–46451 [View Article][PubMed]
    [Google Scholar]
  20. Fuhrer T., Sauer U. 2009; Different biochemical mechanisms ensure network-wide balancing of reducing equivalents in microbial metabolism. J Bacteriol 191:2112–2121 [View Article][PubMed]
    [Google Scholar]
  21. Fuhrer T., Fischer E., Sauer U. 2005; Experimental identification and quantification of glucose metabolism in seven bacterial species. JBacteriol 187:1581–1590 [View Article][PubMed]
    [Google Scholar]
  22. Gerosa L., Kochanowski K., Heinemann M., Sauer U. 2013; Dissecting specific and global transcriptional regulation of bacterial gene expression. Mol Syst Biol 9:658 [View Article][PubMed]
    [Google Scholar]
  23. Gerosa L., Haverkorn van Rijsewijk B. R. B., Christodoulou D., Kochanowski K., Schmidt T. S., Noor E., Sauer U. 2015; Pseudo-transition analysis identifies the key regulators of dynamic metabolic adaptations from steady-state data. Cell Syst 1:270–282 [View Article][PubMed]
    [Google Scholar]
  24. Harold F. 1986 The Vital Force: a Study of Bioenergetics New York, NY: W.H. Freeman and Company;
    [Google Scholar]
  25. Hart B. R., Blumenthal R. M. 2011; Unexpected coregulator range for the global regulator Lrp of Escherichia coli and Proteus mirabilis . J Bacteriol 193:1054–1064 [View Article][PubMed]
    [Google Scholar]
  26. Hart Y., Madar D., Yuan J., Bren A., Mayo A. E., Rabinowitz J. D., Alon U. 2011; Robust control of nitrogen assimilation by a bifunctional enzyme in E. coli . Mol Cell 41:117–127 [View Article][PubMed]
    [Google Scholar]
  27. Haverkorn van Rijsewijk B. R. B., Nanchen A., Nallet S., Kleijn R. J., Sauer U. 2011; Large-scale 13C-flux analysis reveals distinct transcriptional control of respiratory and fermentative metabolism in Escherichia coli . Mol Syst Biol 7:477 [View Article][PubMed]
    [Google Scholar]
  28. Holm A. K., Blank L. M., Oldiges M., Schmid A., Solem C., Jensen P. R., Vemuri G. N. 2010; Metabolic and transcriptional response to cofactor perturbations in Escherichia coli . J Biol Chem 285:17498–17506 [View Article][PubMed]
    [Google Scholar]
  29. Hung S.-P., Baldi P., Hatfield G. W. 2002; Global gene expression profiling in Escherichia coli K12. The effects of leucine-responsive regulatory protein. J Biol Chem 277:40309–40323 [View Article][PubMed]
    [Google Scholar]
  30. Iuchi S., Lin E. C. 1988; arcA (dye), a global regulatory gene in Escherichia coli mediating repression of enzymes in aerobic pathways. Proc Natl Acad Sci USA 85:1888–1892 [View Article]
    [Google Scholar]
  31. Kawai S., Mori S., Mukai T., Hashimoto W., Murata K. 2001; Molecular characterization of Escherichia coli NAD kinase. Eur J Biochem 268:4359–4365 [CrossRef]
    [Google Scholar]
  32. Leavitt R. I., Umbarger H. E. 1962; Isoleucine and valine metabolism in Escherichia coli. XI. K-12: Valine inhibition of the growth of Escherichia coli strain. J Bacteriol 83:624–630[PubMed]
    [Google Scholar]
  33. Liu M., Durfee T., Cabrera J. E., Zhao K., Jin D. J., Blattner F. R. 2005; Global transcriptional programs reveal a carbon source foraging strategy by Escherichia coli . J Biol Chem 280:15921–15927 [View Article][PubMed]
    [Google Scholar]
  34. Michal G. 1999 Biochemical Pathways Heidelberg, Germany: Spektrum Akademischer Verlag;
    [Google Scholar]
  35. Neidhardt F. C., Bott M. 1990 Physiology of the Bacterial Cell: a Molecular Approach Sunderland, MA: Sinauer Associates;
    [Google Scholar]
  36. Outten C. E., Culotta V. C. 2003; A novel NADH kinase is the mitochondrial source of NADPH in Saccharomyces cerevisiae . EMBO J 22:2015–2024 [View Article][PubMed]
    [Google Scholar]
  37. Overkamp K. M., Bakker B. M., Steensma H. Y., van Dijken J. P., Pronk J. T. 2002; Two mechanisms for oxidation of cytosolic NADPH by Kluyveromyces lactis mitochondria. Yeast 19:813–824 [View Article][PubMed]
    [Google Scholar]
  38. Perrenoud A., Sauer U. 2005; Impact of global transcriptional regulation by ArcA, ArcB, Cra, Crp, Cya, Fnr, and Mlc on glucose catabolism in Escherichia coli . J Bacteriol 187:3171–3179 [View Article][PubMed]
    [Google Scholar]
  39. Ralser M., Wamelink M. M., Kowald A., Gerisch B., Heeren G., Struys E. A., Klipp E., Jakobs C., Breitenbach M. et al. 2007; Dynamic rerouting of the carbohydrate flux is key to counteracting oxidative stress. J Biol 6:10 [View Article][PubMed]
    [Google Scholar]
  40. Ralser M., Wamelink M. M. C., Latkolik S., Jansen E. E. W., Lehrach H., Jakobs C. 2009; Metabolic reconfiguration precedes transcriptional regulation in the antioxidant response. Nat Biotechnol 27:604–605 [View Article]
    [Google Scholar]
  41. Ritz D., Beckwith J. 2001; Roles of thiol-redox pathways in bacteria. Annu Rev Microbiol 55:21–48 [View Article][PubMed]
    [Google Scholar]
  42. Ruiz J., Haneburger I., Jung K. 2011; Identification of ArgP and Lrp as transcriptional regulators of lysP, the gene encoding the specific lysine permease of Escherichia coli . J Bacteriol 193:2536–2548 [View Article][PubMed]
    [Google Scholar]
  43. Samorski M., Müller-Newen G., Büchs J. 2005; Quasi-continuous combined scattered light and fluorescence measurements: a novel measurement technique for shaken microtiter plates. Biotechnol Bioeng 92:61–68 [View Article][PubMed]
    [Google Scholar]
  44. Sauer U., Canonaco F., Heri S., Perrenoud A., Fischer E. 2004; The soluble and membrane-bound transhydrogenases UdhA and PntAB have divergent functions in NADPH metabolism of Escherichia coli . J Biol Chem 279:6613–6619 [View Article][PubMed]
    [Google Scholar]
  45. Sherlock G., Hernandez-Boussard T., Kasarskis A., Binkley G., Matese J. C., Dwight S. S., Kaloper M., Weng S., Jin H. et al. 2001; The Stanford Microarray Database. Nucleic Acids Res 29:152–155 [View Article][PubMed]
    [Google Scholar]
  46. Singh R., Lemire J., Mailloux R. J., Appanna V. D. 2008; A novel strategy involved anti-oxidative defense: the conversion of NADH into NADPH by a metabolic network. PLoS One 3:e2682 [View Article][PubMed]
    [Google Scholar]
  47. Smith C. A. 1992; Biosynthesis and fueling. In Physiology of the Bacterial Cell: a Molecular Approach Edited by Neidhardt F. C., LIngraham J., Schaechter M. Sunderland, MA: Sinauer Associates;
    [Google Scholar]
  48. Verho R., Richard P., Jonson P. H., Sundqvist L., Londesborough J., Penttilä M. 2002; Identification of the first fungal NADP-GAPDH from Kluyveromyces lactis . Biochemistry 41:13833–13838[PubMed] [CrossRef]
    [Google Scholar]
  49. Zamboni N., Fischer E., Laudert D., Aymerich S., Hohmann H. P., Sauer U. 2004; The Bacillus subtilis yqjI gene encodes the NADP+-dependent 6-P-gluconate dehydrogenase in the pentose phosphate pathway. J Bacteriol 186:4528–4534 [View Article][PubMed]
    [Google Scholar]
  50. Zaslaver A., Mayo A. E., Rosenberg R., Bashkin P., Sberro H., Tsalyuk M., Surette M. G., Alon U. 2004; Just-in-time transcription program in metabolic pathways. Nat Genet 36:486–491 [View Article][PubMed]
    [Google Scholar]
  51. Zaslaver A., Bren A., Ronen M., Itzkovitz S., Kikoin I., Shavit S., Liebermeister W., Surette M. G., Alon U. 2006; A comprehensive library of fluorescent transcriptional reporters for Escherichia coli . Nat Methods 3:623–628 [View Article]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.000346
Loading
/content/journal/micro/10.1099/mic.0.000346
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