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Archives of Microbiology

, Volume 192, Issue 9, pp 691–702 | Cite as

The small RNA Aar in Acinetobacter baylyi: a putative regulator of amino acid metabolism

  • Dominik Schilling
  • Sven Findeiß
  • Andreas S. Richter
  • Jennifer A. Taylor
  • Ulrike Gerischer
Original Paper

Abstract

Small non-coding RNAs (sRNAs) are key players in prokaryotic metabolic circuits, allowing the cell to adapt to changing environmental conditions. Regulatory interference by sRNAs in cellular metabolism is often facilitated by the Sm-like protein Hfq. A search for novel sRNAs in A. baylyi intergenic regions was performed by a biocomputational screening. One candidate, Aar, encoded between trpS and sucD showed Hfq dependency in Northern blot analysis. Aar was expressed strongly during stationary growth phase in minimal medium; in contrast, in complex medium, strongest expression was in the exponential growth phase. Whereas over-expression of Aar in trans did not affect bacterial growth, seven mRNA targets predicted by two in silico approaches were upregulated in stationary growth phase. All seven mRNAs are involved in A. baylyi amino acid metabolism. A putative binding site for Lrp, the global regulator of branched-chain amino acids in E. coli, was observed within the aar gene. Both facts imply an Aar participation in amino acid metabolism.

Keywords

A. baylyi Hfq IntaRNA RNAup sRNAs 

Notes

Acknowledgments

We would like to thank Björn Voss for his predictions of sRNA genes in Acinetobacter and Véronique de Berardinis for her effort to delete aar. Furthermore, we would like to thank Iris Steiner for her contribution. This work was supported by the German Federal Ministry of Education and Research (BMBF grant 0313921 FRISYS to A.S.R.); the German Research Foundation (DFG grant SPP1258 to S.F. STA850/7-1 and A.S.R. BA2168/2-1); and the state of Baden-Württemberg, Germany (personal LGFG grant to D.S.).

References

  1. Altuvia S (2007) Identification of bacterial small non-coding RNAs: experimental approaches. Curr Opin Microbiol 10:257–261CrossRefPubMedGoogle Scholar
  2. Argaman L, Hershberg R, Vogel J, Bejerano G, Wagner EG, Margalit H, Altuvia S (2001) Novel small RNA-encoding genes in the intergenic regions of Escherichia coli. Curr Biol 11:941–950CrossRefPubMedGoogle Scholar
  3. Axmann IM, Kensche P, Vogel J, Kohl S, Herzel H, Hess WR (2005) Identification of cyanobacterial non-coding RNAs by comparative genome analysis. Genome Biol 6:R73CrossRefPubMedGoogle Scholar
  4. Barbe V, Vallenet D, Fonknechten N, Kreimeyer A, Oztas S, Labarre L, Cruveiller S, Robert C, Duprat S, Wincker P, Ornston LN, Weissenbach J, Marliere P, Cohen GN, Medigue C (2004) Unique features revealed by the genome sequence of Acinetobacter sp. ADP1, a versatile and naturally transformation competent bacterium. Nucleic Acids Res 32:5766–5779CrossRefPubMedGoogle Scholar
  5. Bejerano-Sagie M, Xavier KB (2007) The role of small RNAs in quorum sensing. Curr Opin Microbiol 10:189–198CrossRefPubMedGoogle Scholar
  6. Boisset S, Geissmann T, Huntzinger E, Fechter P, Bendridi N, Possedko M, Chevalier C, Helfer AC, Benito Y, Jacquier A, Gaspin C, Vandenesch F, Romby P (2007) Staphylococcus aureus RNAIII coordinately represses the synthesis of virulence factors and the transcription regulator Rot by an antisense mechanism. Genes Dev 21:1353–1366CrossRefPubMedGoogle Scholar
  7. Brescia CC, Mikulecky PJ, Feig AL, Sledjeski DD (2003) Identification of the Hfq-binding site on DsrA RNA: Hfq binds without altering DsrA secondary structure. Rna 9:33–43CrossRefPubMedGoogle Scholar
  8. Buck D, Guest JR (1989) Overexpression and site-directed mutagenesis of the succinyl-CoA synthetase of Escherichia coli and nucleotide sequence of a gene (g30) that is adjacent to the suc operon. Biochem J 260:737–747PubMedGoogle Scholar
  9. Busch A, Richter AS, Backofen R (2008) IntaRNA: efficient prediction of bacterial sRNA targets incorporating target site accessibility and seed regions. Bioinformatics 24:2849–2856CrossRefPubMedGoogle Scholar
  10. Calvo JM, Matthews RG (1994) The leucine-responsive regulatory protein, a global regulator of metabolism in Escherichia coli. Microbiol Rev 58:466–490PubMedGoogle Scholar
  11. Cui Y, Wang Q, Stormo GD, Calvo JM (1995) A consensus sequence for binding of Lrp to DNA. J Bacteriol 177:4872–4880PubMedGoogle Scholar
  12. de Berardinis V, Vallenet D, Castelli V, Besnard M, Pinet A, Cruaud C, Samair S, Lechaplais C, Gyapay G, Richez C, Durot M, Kreimeyer A, Le Fevre F, Schachter V, Pezo V, Doring V, Scarpelli C, Medigue C, Cohen GN, Marliere P, Salanoubat M, Weissenbach J (2008) A complete collection of single-gene deletion mutants of Acinetobacter baylyi ADP1. Mol Syst Biol 4:174PubMedGoogle Scholar
  13. de Boer PA, Crossley RE, Rothfield LI (1989) A division inhibitor and a topological specificity factor coded for by the minicell locus determine proper placement of the division septum in E. coli. Cell 56:641–649PubMedGoogle Scholar
  14. Delihas N, Forst S (2001) MicF: an antisense RNA gene involved in response of Escherichia coli to global stress factors. J Mol Biol 313:1–12CrossRefPubMedGoogle Scholar
  15. Figurski DH, Helinski DR (1979) Replication of an origin-containing derivative of plasmid RK2 dependent on a plasmid function provided in trans. Proc Natl Acad Sci USA 76:1648–1652CrossRefPubMedGoogle Scholar
  16. Folichon M, Allemand F, Regnier P, Hajnsdorf E (2005) Stimulation of poly(A) synthesis by Escherichia coli poly(A)polymerase I is correlated with Hfq binding to poly(A) tails. FEBS J 272:454–463CrossRefPubMedGoogle Scholar
  17. Forner J, Weber B, Thuss S, Wildum S, Binder S (2007) Mapping of mitochondrial mRNA termini in Arabidopsis thaliana: t-elements contribute to 5′ and 3′ end formation. Nucleic Acids Res 35:3676–3692CrossRefPubMedGoogle Scholar
  18. Gardner PP, Daub J, Tate JG, Nawrocki EP, Kolbe DL, Lindgreen S, Wilkinson AC, Finn RD, Griffiths-Jones S, Eddy SR, Bateman A (2009) Rfam: updates to the RNA families database. Nucleic Acids Res 37:D136–D140CrossRefPubMedGoogle Scholar
  19. Geissmann TA, Touati D (2004) Hfq, a new chaperoning role: binding to messenger RNA determines access for small RNA regulator. EMBO J 23:396–405CrossRefPubMedGoogle Scholar
  20. Gorke B, Vogel J (2008) Non-coding RNA control of the making and breaking of sugars. Genes Dev 22:2914–2925CrossRefPubMedGoogle Scholar
  21. Gottesman S (2005) Micros for microbes: non-coding regulatory RNAs in bacteria. Trends Genet 21:399–404CrossRefPubMedGoogle Scholar
  22. Hajnsdorf E, Regnier P (2000) Host factor Hfq of Escherichia coli stimulates elongation of poly(A) tails by poly(A) polymerase I. Proc Natl Acad Sci USA 97:1501–1505CrossRefPubMedGoogle Scholar
  23. Hanahan D (1983) Studies on transformation of Escherichia coli with plasmids. J Mol Biol 166:557–580CrossRefPubMedGoogle Scholar
  24. Hershberg R, Altuvia S, Margalit H (2003) A survey of small RNA-encoding genes in Escherichia coli. Nucleic Acids Res 31:1813–1820CrossRefPubMedGoogle Scholar
  25. Kalamorz F, Reichenbach B, Marz W, Rak B, Gorke B (2007) Feedback control of glucosamine-6-phosphate synthase GlmS expression depends on the small RNA GlmZ and involves the novel protein YhbJ in Escherichia coli. Mol Microbiol 65:1518–1533CrossRefPubMedGoogle Scholar
  26. Kawamoto H, Koide Y, Morita T, Aiba H (2006) Base-pairing requirement for RNA silencing by a bacterial small RNA and acceleration of duplex formation by Hfq. Mol Microbiol 61:1013–1022CrossRefPubMedGoogle Scholar
  27. Kawano M, Reynolds AA, Miranda-Rios J, Storz G (2005) Detection of 5′- and 3′-UTR-derived small RNAs and cis-encoded antisense RNAs in Escherichia coli. Nucleic Acids Res 33:1040–1050CrossRefPubMedGoogle Scholar
  28. Keen NT, Tamaki S, Kobayashi D, Trollinger D (1988) Improved broad-host-range plasmids for DNA cloning in Gram-negative bacteria. Gene 70:191–197CrossRefPubMedGoogle Scholar
  29. Kingsford CL, Ayanbule K, Salzberg SL (2007) Rapid, accurate, computational discovery of Rho-independent transcription terminators illuminates their relationship to DNA uptake. Genome Biol 8:R22CrossRefPubMedGoogle Scholar
  30. Le Derout J, Folichon M, Briani F, Deho G, Regnier P, Hajnsdorf E (2003) Hfq affects the length and the frequency of short oligo(A) tails at the 3′ end of Escherichia coli rpsO mRNAs. Nucleic Acids Res 31:4017–4023CrossRefPubMedGoogle Scholar
  31. Le Derout J, Boni IV, Regnier P, Hajnsdorf E (2010) Hfq affects mRNA levels independently of degradation. BMC Mol Biol 11:17CrossRefPubMedGoogle Scholar
  32. Lin R, D’Ari R, Newman EB (1992) Lambda placMu insertions in genes of the leucine regulon: extension of the regulon to genes not regulated by leucine. J Bacteriol 174:1948–1955PubMedGoogle Scholar
  33. Liu MY, Gui G, Wei B, Preston JF 3rd, Oakford L, Yuksel U, Giedroc DP, Romeo T (1997) The RNA molecule CsrB binds to the global regulatory protein CsrA and antagonizes its activity in Escherichia coli. J Biol Chem 272:17502–17510CrossRefPubMedGoogle Scholar
  34. Livny J, Waldor MK (2007) Identification of small RNAs in diverse bacterial species. Curr Opin Microbiol 10:96–101CrossRefPubMedGoogle Scholar
  35. Livny J, Brencic A, Lory S, Waldor MK (2006) Identification of 17 Pseudomonas aeruginosa sRNAs and prediction of sRNA-encoding genes in 10 diverse pathogens using the bioinformatic tool sRNAPredict2. Nucleic Acids Res 34:3484–3493CrossRefPubMedGoogle Scholar
  36. Majdalani N, Cunning C, Sledjeski D, Elliott T, Gottesman S (1998) DsrA RNA regulates translation of RpoS message by an anti-antisense mechanism, independent of its action as an antisilencer of transcription. Proc Natl Acad Sci USA 95:12462–12467CrossRefPubMedGoogle Scholar
  37. Maki K, Uno K, Morita T, Aiba H (2008) RNA, but not protein partners, is directly responsible for translational silencing by a bacterial Hfq-binding small RNA. Proc Natl Acad Sci USA 105:10332–10337CrossRefPubMedGoogle Scholar
  38. Masse E, Gottesman S (2002) A small RNA regulates the expression of genes involved in iron metabolism in Escherichia coli. Proc Natl Acad Sci USA 99:4620–4625CrossRefPubMedGoogle Scholar
  39. Masse E, Escorcia FE, Gottesman S (2003) Coupled degradation of a small regulatory RNA and its mRNA targets in Escherichia coli. Genes Dev 17:2374–2383CrossRefPubMedGoogle Scholar
  40. Masse E, Salvail H, Desnoyers G, Arguin M (2007) Small RNAs controlling iron metabolism. Curr Opin Microbiol 10:140–145CrossRefPubMedGoogle Scholar
  41. Mizuno T, Chou MY, Inouye M (1984) A unique mechanism regulating gene expression: translational inhibition by a complementary RNA transcript (micRNA). Proc Natl Acad Sci USA 81:1966–1970CrossRefPubMedGoogle Scholar
  42. Moll I, Afonyushkin T, Vytvytska O, Kaberdin VR, Blasi U (2003) Coincident Hfq binding and RNase E cleavage sites on mRNA and small regulatory RNAs. Rna 9:1308–1314CrossRefPubMedGoogle Scholar
  43. Mückstein U, Tafer H, Hackermuller J, Bernhart SH, Stadler PF, Hofacker IL (2006) Thermodynamics of RNA-RNA binding. Bioinformatics 22:1177–1182CrossRefPubMedGoogle Scholar
  44. Munch R, Hiller K, Barg H, Heldt D, Linz S, Wingender E, Jahn D (2003) PRODORIC: prokaryotic database of gene regulation. Nucleic Acids Res 31:266–269CrossRefPubMedGoogle Scholar
  45. Newman EB, Lin R (1995) Leucine-responsive regulatory protein: a global regulator of gene expression in E. coli. Annu Rev Microbiol 49:747–775CrossRefPubMedGoogle Scholar
  46. Oelmüller U, Krüger N, Steinbüchel A, Freidrich CG (1990) Isolation of prokaryotic RNA and detection of specific mRNA with biotinylated probes. J Microbiol Methods 11:73–81CrossRefGoogle Scholar
  47. Pfeiffer V, Sittka A, Tomer R, Tedin K, Brinkmann V, Vogel J (2007) A small non-coding RNA of the invasion gene island (SPI-1) represses outer membrane protein synthesis from the Salmonella core genome. Mol Microbiol 66:1174–1191CrossRefPubMedGoogle Scholar
  48. Regnier P, Hajnsdorf E (2008) The role of RNA chaperone Hfq in poly(A) metabolism methods to determine positions, abundance, and lengths of short oligo(A) tails. Methods Enzymol 447:161–181CrossRefPubMedGoogle Scholar
  49. Sambrook J, Russel DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory Press, Cold Spring HarborGoogle Scholar
  50. Schilling D, Gerischer U (2009) The Acinetobacter baylyi hfq gene encodes a large protein with an unusual C terminus. J Bacteriol 191:5553–5562CrossRefPubMedGoogle Scholar
  51. Schimmel PR, Soll D (1979) Aminoacyl-tRNA synthetases: general features and recognition of transfer RNAs. Annu Rev Biochem 48:601–648CrossRefPubMedGoogle Scholar
  52. Sharma CM, Darfeuille F, Plantinga TH, Vogel J (2007) A small RNA regulates multiple ABC transporter mRNAs by targeting C/A-rich elements inside and upstream of ribosome-binding sites. Genes Dev 21:2804–2817CrossRefPubMedGoogle Scholar
  53. Sonnleitner E, Sorger-Domenigg T, Madej MJ, Findeiss S, Hackermuller J, Huttenhofer A, Stadler PF, Blasi U, Moll I (2008) Detection of small RNAs in Pseudomonas aeruginosa by RNomics and structure-based bioinformatic tools. Microbiology 154:3175–3187CrossRefPubMedGoogle Scholar
  54. Soper TJ, Woodson SA (2008) The rpoS mRNA leader recruits Hfq to facilitate annealing with DsrA sRNA. Rna 14:1907–1917CrossRefPubMedGoogle Scholar
  55. Starmer J, Stomp A, Vouk M, Bitzer D (2006) Predicting Shine-Dalgarno sequence locations exposes genome annotation errors. PLoS Comput Biol 2:e57CrossRefPubMedGoogle Scholar
  56. Trautwein G, Gerischer U (2001) Effects exerted by transcriptional regulator PcaU from Acinetobacter sp. strain ADP1. J Bacteriol 183:873–881CrossRefPubMedGoogle Scholar
  57. Tu KC, Bassler BL (2007) Multiple small RNAs act additively to integrate sensory information and control quorum sensing in Vibrio harveyi. Genes Dev 21:221–233CrossRefPubMedGoogle Scholar
  58. Udekwu KI, Darfeuille F, Vogel J, Reimegard J, Holmqvist E, Wagner EG (2005) Hfq-dependent regulation of OmpA synthesis is mediated by an antisense RNA. Genes Dev 19:2355–2366CrossRefPubMedGoogle Scholar
  59. Valentin-Hansen P, Eriksen M, Udesen C (2004) The bacterial Sm-like protein Hfq: a key player in RNA transactions. Mol Microbiol 51:1525–1533CrossRefPubMedGoogle Scholar
  60. Vanderpool CK (2007) Physiological consequences of small RNA-mediated regulation of glucose-phosphate stress. Curr Opin Microbiol 10:146–151CrossRefPubMedGoogle Scholar
  61. Vaneechoutte M, Young DM, Ornston LN, De Baere T, Nemec A, Van Der Reijden T, Carr E, Tjernberg I, Dijkshoorn L (2006) Naturally transformable Acinetobacter sp. strain ADP1 belongs to the newly described species Acinetobacter baylyi. Appl Environ Microbiol 72:932–936CrossRefPubMedGoogle Scholar
  62. Vogel J, Papenfort K (2006) Small non-coding RNAs and the bacterial outer membrane. Curr Opin Microbiol 9:605–611CrossRefPubMedGoogle Scholar
  63. Voss B, Georg J, Schon V, Ude S, Hess WR (2009) Biocomputational prediction of non-coding RNAs in model cyanobacteria. BMC Genomics 10:123CrossRefPubMedGoogle Scholar
  64. Washietl S, Hofacker IL, Stadler PF (2005) Fast and reliable prediction of non-coding RNAs. Proc Natl Acad Sci USA 102:2454–2459CrossRefPubMedGoogle Scholar
  65. Wassarman KM (2007) 6S RNA: a small RNA regulator of transcription. Curr Opin Microbiol 10:164–168CrossRefPubMedGoogle Scholar
  66. Willkomm DK, Minnerup J, Hüttenhofer A, Hartmann RK (2005) Experimental RNomics in Aquifex aeolicus: identification of small non-coding RNAs and the putative 6S RNA homolog. Nucleic Acids Res 33:1949–1960CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Dominik Schilling
    • 1
    • 5
  • Sven Findeiß
    • 2
  • Andreas S. Richter
    • 3
  • Jennifer A. Taylor
    • 4
  • Ulrike Gerischer
    • 1
  1. 1.Institute of Microbiology and BiotechnologyUniversity of UlmUlmGermany
  2. 2.Bioinformatics Group, Department of Computer Science, Interdisciplinary Center for BioinformaticsUniversity of LeipzigLeipzigGermany
  3. 3.Bioinformatics Group, Department of Computer ScienceUniversity of FreiburgFreiburgGermany
  4. 4.Department of MicrobiologyUniversity of GeorgiaAthensUSA
  5. 5.Department of Conservative Dentistry and PeriodontologyUniversity of UlmUlmGermany

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