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Subcellular localization and tissue specific expression of amidase 1 from Arabidopsis thaliana

Planta volume 224, pages 1241–1253 (2006)Cite this article

Abstract

Amidase 1 (AMI1) from Arabidopsis thaliana converts indole-3-acetamide (IAM), into indole-3-acetic acid (IAA). AMI1 is part of a small isogene family comprising seven members in A. thaliana encoding proteins which share a conserved glycine- and serine-rich amidase-signature. One member of this family has been characterized as an N-acylethanolamine-cleaving fatty acid amidohydrolase (FAAH) and two other members are part of the preprotein translocon of the outer envelope of chloroplasts (Toc complex) or mitochondria (Tom complex) and presumably lack enzymatic activity. Among the hitherto characterized proteins of this family, AMI1 is the only member with indole-3-acetamide hydrolase activity, and IAM is the preferred substrate while N-acylethanolamines and oleamide are not hydrolyzed significantly, thus suggesting a role of AMI1 in auxin biosynthesis. Whereas the enzymatic function of AMI1 has been determined in vitro, the subcellular localization of the enzyme remained unclear. By using different GFP-fusion constructs and an A. thaliana transient expression system, we show a cytoplasmic localization of AMI1. In addition, RT-PCR and anti-amidase antisera were used to examine tissue specific expression of AMI1 at the transcriptional and translational level, respectively. AMI1-expression is strongest in places of highest IAA content in the plant. Thus, it is concluded that AMI1 may be involved in de novo IAA synthesis in A. thaliana.

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Abbreviations

DsRed:

Red fluorescent protein from Discosoma sp.

GFP:

Green fluorescent protein

IAA:

Indole-3-acetic acid

IAM:

Indole-3-acetamide

NAA:

1-Naphthaleneacetic acid

NAE:

N-acylethanolamine

NAM:

1-Naphthaleneacetamide

References

  • An YQ, McDowell JM, Huang S, McKinney EC, Chambliss S, Meagher RB (1996) Strong, constitutive expression of the Arabidopsis ACT2/ACT8 actin subclass in vegetative tissues. Plant J 10:107–121

    Article  PubMed  Google Scholar 

  • Arai Y, Kawaguchi M, Syono K, Ikuta A (2004) Partial purification of an enzyme hydrolyzing indole-3-acetamide from rice cells. J Plant Res 117:191–198

    Article  PubMed  Google Scholar 

  • Ausubel FA, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K (2000) Current protocols in molecular biology. Wiley, New York

    Google Scholar 

  • Bracey MH, Hanson MA, Masuda KR, Stevens RC, Cravatt BF (2002) Structural adaptations in a membrane enzyme that terminates endocannabinoid signaling. Science 298:1793–1796

    Article  PubMed  Google Scholar 

  • Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  PubMed  Google Scholar 

  • Brenner S, Johnson M, Bridgham J, Golda G, Lloyd DH, Johnson D, Luo S, McCurdy S, Foy M, Ewan M, Roth R, George D, Eletr S, Albrecht G, Vermaas E, Williams SR, Moon K, Burcham T, Pallas M, DuBridge RB, Kirchner J, Fearon K, Mao J, Corcoran K (2000) Gene expression analysis by massively parallel signature sequencing (MPSS) on microbead arrays. Nat Biotechnol 18:630–634

    Article  PubMed  Google Scholar 

  • Bruce WB, Christensen AH, Klein T, Fromm M, Quail PH (1989) Photoregulation of a phytochrome gene promotor from oat transferred into rice by particle bombardment. Proc Natl Acad Sci USA 86:9692–9696

    Article  PubMed  Google Scholar 

  • Chew O, Lister R, Qbadou S, Heazlewood JL, Soll J, Schleiff E, Millar AH, Whelan J (2004) A plant outer mitochondrial membrane protein with high amino acid sequence identity to a chloroplast protein import receptor. FEBS Lett 557:109–114

    Article  PubMed  Google Scholar 

  • Cravatt BF, Giang DK, Mayfield SP, Boger DL, Lerner RA, Gilula NB (1996) Molecular characterization of an enzyme that degrades neuromodulatory fatty-acid amides. Nature 384:83–87

    Article  PubMed  Google Scholar 

  • Cutler SR, Somerville CR (2005) Imaging plant cell death: GFP-Nit1 aggregation marks an early step of wound and herbicide induced cell death. BMC Plant Biol 5:4

    Article  PubMed  Google Scholar 

  • Finer JJ, Vain P, Jones MW, McMullen MD (1992) Development of the particle inflow gun for DNA delivery to plant cells. Plant Cell Rep 11:323–328

    Article  Google Scholar 

  • Giang DK, Cravatt BF (1997) Molecular characterization of human and mouse fatty acid amide hydrolases. Proc Natl Acad Sci USA 94:2238–2242

    Article  PubMed  Google Scholar 

  • Haseloff J, Siemering KR, Prasher DC, Hodge S (1997) Removal of a cryptic intron and subcellular localization of green fluorescent protein are required to mark transgenic Arabidopsis plants brightly. Proc Natl Acad Sci USA 94:2122–2127

    Article  PubMed  Google Scholar 

  • Hofmann NR, Theg SM (2004) Physcomitrella patens as a model for the study of chloroplast protein transport: conserved machineries between vascular and non-vascular plants. Plant Mol Biol 53:621–632

    Google Scholar 

  • Kawaguchi M, Kobayashi M, Sakurai A, Syono K (1991) The presence of an enzyme that converts indole-3-acetamide into IAA in wild and cultivated rice. Plant Cell Physiol 32:143–149

    Google Scholar 

  • Kawaguchi M, Fujioka S, Sakurai A, Yamaki YT, Syono K (1993) Presence of a pathway for the biosynthesis of auxin via indole-3-acetamide in trifoliata orange. Plant Cell Physiol 34:121–128

    Google Scholar 

  • Kemper E, Waffenschmidt S, Weiler EW, Rausch T, Schröder J (1985) T-DNA coded auxin formation in crown gall cells. Planta 163:257–262

    Article  Google Scholar 

  • Kim YS, Kang SW (1994) Novel malonamidases in Bradyrhizobium japonicum. Purification, characterization, and immunological comparison. J Biol Chem 269:8014–8021

    PubMed  Google Scholar 

  • Kleiner O, Kircher S, Harter K, Batschauer A (1999) Nuclear localization of the Arabidopsis blue light receptor cryptochrome 2. Plant J 19:289–296

    Article  PubMed  Google Scholar 

  • Kobayashi M, Komeda H, Nagasawa T, Nishiyama M, Horinouchi S, Beppu T, Yamada H, Shimizu S (1993) Amidase coupled with low-molecular-mass nitrile hydratase from Rhodococcus rhodochrous J1. Sequencing and expression of the gene and purification and characterization of the gene product. Eur J Biochem 217:327–336

    Article  PubMed  Google Scholar 

  • Lee YJ, Sohn EJ, Lee KH, Lee DW, Hwang I (2004) The transmembrane domain of AtToc64 and its C-terminal lysine-rich flanking region are targeting signals to the chloroplast outer envelope membrane. Mol Cells 17:281–291

    PubMed  Google Scholar 

  • Madera M, Vogel C, Kummerfeld SK, Chothia C, Gough J (2004) The SUPERFAMILY database in 2004: additions and improvements. Nucleic Acids Res 32:D235–D239

    Article  PubMed  Google Scholar 

  • Mayaux J-F, Cerbelaud E, Soubrier F, Faucher D, Pétré D (1990) Purification, cloning and primary structure of an enantiomer-selective amidase from Brevibacterium sp. strain R312: evidence for genetic coupling with nitrile hydrolase. J Bacteriol 172:6764–6773

    PubMed  Google Scholar 

  • Metz AM, Timmer RT, Browning KS (1992) Sequences for two cDNAs encoding Arabidopsis thaliana eukaryotic protein synthesis initiation factor 4A. Gene 120:313–314

    Article  PubMed  Google Scholar 

  • Müller A, Weiler EW (2000) IAA-synthase, an enzyme complex from Arabidopsis thaliana catalyzing the formation of indole-3-acetic acid from (S)-tryptophan. Biol Chem 381:679–686

    Article  PubMed  Google Scholar 

  • Müller A, Düchting P, Weiler EW (2002) A multiplex GC–MS/MS technique for the sensitive and quantitative single-run analysis of acidic phytohormones and related compounds, and its application to Arabidopsis thaliana. Planta 216:44–56

    Article  PubMed  Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497

    Article  Google Scholar 

  • Patricelli MP, Cravatt BF (2000) Clarifying the catalytic roles of conserved residues in the amidase signature family. J Biol Chem 275:19177–19184

    Article  PubMed  Google Scholar 

  • Pollmann S, Müller A, Piotrowski M, Weiler EW (2002) Occurrence and formation of indole-3-acetamide in Arabidopsis thaliana. Planta 216:155–161

    Article  PubMed  Google Scholar 

  • Pollmann S, Neu D, Weiler EW (2003) Molecular cloning and characterization of an amidase from Arabidopsis thaliana capable of converting indole-3-acetamide into the plant growth hormone, indole-3-acetic acid. Phytochemistry 62:293–300

    Article  PubMed  Google Scholar 

  • Reddy VS, Ali GS, Reddy AS (2002) Genes encoding calmodulin-binding proteins in the Arabidopsis genome. J Biol Chem 277:9840–9852

    Article  PubMed  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor

    Google Scholar 

  • Saotome M, Shirahata K, Nishimura R, Yakaba M, Kawaguchi M, Syono K, Kitsuwa T, Ishii Y, Nakamura T (1993) The identification of indole-3-acetic acid and indole-3-acetamide in the hypocotyls of Japanese cherry. Plant Cell Physiol 34:157–159

    Google Scholar 

  • Saslowsky DE, Warek U, Winkel BSJ (2005) Nuclear localization of flavonoid enzymes in Arabidopsis. J Biol Chem 280:23735–23740

    Article  PubMed  Google Scholar 

  • Scharf KD, Nover L (1982) Heat-shock-induced alterations of ribosomal protein phosphorylation in plant cell cultures. Cell Sep 30:427–437

    Google Scholar 

  • Shin S, Lee TH, Ha NC, Koo HM, Kim SY, Lee HS, Kim YS, Oh BH (2002) Structure of malonamidase E2 reveals a novel Ser-cisSer-Lys catalytic triad in a new serine hydrolase fold that is prevalent in nature. EMBO J 21:2509–2516

    Article  PubMed  Google Scholar 

  • Shin S, Yun YS, Koo YS, Kim YS, Choi KY, Oh BH (2003) Characterization of a novel Ser-cisSer-Lys catalytic triad in comparison with the classical Ser-His-Asp triad. J Biol Chem 278:24937–24943

    Article  PubMed  Google Scholar 

  • Shrestha R, Noordermeer MA, Van der Stelt M, Veldink GA, Chapman KD (2002) N-acylethanolamines are metabolized by lipoxygenase and amidohydrolase in competing pathways during cottonseed imbibition. Plant Physiol 130:391–401

    Article  PubMed  Google Scholar 

  • Shrestha R, Dixon RA, Chapman KD (2003) Molecular identification of a functional homologue of the mammalian fatty acid amide hydrolase in Arabidopsis thaliana. J Biol Chem 278:34990–34997

    Article  PubMed  Google Scholar 

  • Sohrt K, Soll J (2000) Toc64, a new component of the protein translocon of chloroplasts. J Cell Biol 148:1213–1221

    Article  PubMed  Google Scholar 

  • Stelmach BA, Müller A, Hennig P, Gebhardt S, Schubert-Zsilavecz M, Weiler EW (2001) A novel class of oxylipins, sn1-O-(12-oxophytodienoyl)-sn2-O-(hexadecatrienoyl)-monogalactosyl diglyceride, from Arabidopsis thaliana. J Biol Chem 276:12832–12838

    Article  PubMed  Google Scholar 

  • Towbin H, Staehlin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: Procedure and some applications. Proc Natl Acad Sci USA 76:4350–4354

    Article  PubMed  Google Scholar 

  • Upadhyaya NM, Zhou XR, Wu L, Ramm K, Dennis ES (2000) The tms gene as a negative selection marker in rice. Plant Mol Biol Rep 18:227–233

    Google Scholar 

  • Vojta A, Alavi M, Becker T, Hormann F, Kuchler M, Soll J, Thomson R, Schleiff E (2004) The protein translocon of the plastid envelopes. J Biol Chem 279:21401–21405

    Article  PubMed  Google Scholar 

  • Vorwerk S, Biernacki S, Hillebrand H, Janzik I, Müller A, Weiler EW, Piotrowski M (2001) Enzymatic characterization of the recombinant Arabidopsis thaliana nitrilase subfamily encoded by the NIT2/NIT1/NIT3-gene cluster. Planta 212:508–516

    Article  PubMed  Google Scholar 

  • Wachter A, Wolf S, Steininger H, Bogs J, Rausch T (2005) Differential targeting of GSH1 and GSH2 is achieved by multiple transcription initiation: implications for the compartmentation of glutathione biosynthesis in the Brassicaceae. Plant J 41:15–30

    Article  PubMed  Google Scholar 

  • Weiler EW, Schröder J (1987) Hormone genes and the crown gall disease. Trends Biochem Sci 12:271–275

    Article  Google Scholar 

  • Woodward AW, Bartel B (2005) Auxin: regulation, action and interaction. Ann Bot 95:707–735

    Article  PubMed  Google Scholar 

Download references

Acknowledgements

We thank Dr. Rhidaya Shrestha and Dr. Kent D. Chapman (Department of Biological Sciences, Division of Biochemistry and Molecular Biology, University of North Texas, Denton, Texas, USA) for providing the (His)6-tagged FAAH clone. Furthermore, the authors thank Dr. Andreas Wachter (Institute of Plant Sciences, Ruprecht-Karls-University, Heidelberg, Germany) for kindly providing the vectors pFF19-GFP and pFF19-RFP. This work was supported by the Deutsche Forschungsgemeinschaft, Bonn, Germany (Schwerpunktprogramm ‘Molecular Mechanisms of Phytohormone Action’, and subsequently Sonderforschungsbereich 480 ‘Molekulare Biologie komplexer Leistungen von botanischen Systemen’).

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Authors and Affiliations

  1. Department of Plant Physiology, Ruhr-University Bochum, Universitaetsstraße 150, ND 2/27, 44801, Bochum, Germany

    Stephan Pollmann, Daniel Neu, Thomas Lehmann & Elmar W. Weiler

  2. Institut für Anatomie, Universitätsklinikum Essen, Hufelandstraße 55, 45122, Essen, Germany

    Tina Schäfer

  3. The Australian National University, Research School of Biological Sciences (RSBS), Enviromental Biology Group, GPO Box 475, Canberra, ACT, 0200, Australia

    Oliver Berkowitz

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  1. Stephan Pollmann
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  2. Daniel Neu
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  4. Oliver Berkowitz
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  5. Tina Schäfer
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  6. Elmar W. Weiler
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Correspondence to Stephan Pollmann.

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Pollmann, S., Neu, D., Lehmann, T. et al. Subcellular localization and tissue specific expression of amidase 1 from Arabidopsis thaliana . Planta 224, 1241–1253 (2006). https://doi.org/10.1007/s00425-006-0304-2

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Keywords

  • Amidase
  • Amidohydrolase
  • Arabidopsis
  • Brassicaceae
  • Fatty acid amide hydrolase
  • Indole-3-acetic acid
  • Indole-3-acetamide
  • N-acylethanolamine
  • Oleamide