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Identification of a Saxitoxin Biosynthesis Gene with a History of Frequent Horizontal Gene Transfers

Journal of Molecular Evolution volume 67, pages 526–538 (2008)Cite this article

An Erratum to this article was published on 21 February 2009

Abstract

The paralytic shellfish poisoning (PSP) toxins, saxitoxin, and its derivatives, are produced by a complex and unique biosynthetic pathway. It involves reactions that are rare in other metabolic pathways, however, distantly related organisms, such as dinoflagellates and cyanobacteria, produce these toxins by an identical pathway. Speculative explanations for the unusual phylogenetic distribution of this metabolic pathway have been proposed, including a polyphyletic origin, the involvement of symbiotic bacteria, and horizontal gene transfer. This study describes for the first time the identity of one gene, sxt1, that is involved in the biosynthesis of saxitoxin in cyanobacteria. It encoded an O-carbamoyltransferase (OCTASE) that was proposed to carbamoylate the hydroxymethyl side chain of saxitoxin precursor. Orthologues of sxt1 were exclusively present in PSP-toxic strains of cyanobacteria and had a high sequence similarity to each other. L. wollei had a naturally mutated sxt1 gene that encoded an inactive enzyme, and was incapable of producing carbamoylated PSP-toxin analogues, supporting the proposed function of Sxt1. Phylogenetic analysis revealed that OCATSE genes were present exclusively in prokaryotic organisms and were characterized by a high rate of horizontal gene transfer. OCTASE has most likely evolved from an ancestral O-sialoglycoprotein endopeptidase from proteobacteria, whereas the most likely phylogenetic origin of sxt1 was an ancestral α-proteobacterium. The phylogeny of sxt1 suggested that the entire set of genes required for saxitoxin biosynthesis may spread by horizontal gene transfer.

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References

  • Aravind L, Koonin EV (1999) Gleaning non-trivial structural, functional and evolutionary information about proteins by iterative database searches. J Mol Biol 287:1023–1040

    PubMed  Article  CAS  Google Scholar 

  • Beitz E (2000) TeXshade: shading and labeling of multiple sequence alignments using LaTeX2e. Bioinformatics 16:135–139

    PubMed  Article  CAS  Google Scholar 

  • Beltran EC, Neilan BA (2000) Geographical segregation of the neurotoxin-producing cyanobacterium Anabaena circinalis. Appl Environ Microbiol 66:4468–4474

    PubMed  Article  CAS  Google Scholar 

  • Brewer S, Taylor P, Turner M (1980) An adenosine triphosphate-dependent carbamoylphosphate-3-hydroxymethylcephem O-carbamoyltransferase from Streptomyces clavuligerus. Biochem J 185:555–564

    PubMed  CAS  Google Scholar 

  • Coque JJ, Perez-Llarena FJ, Enguita FJ, Fuente JL, Martin JF, Liras P (1995) Characterization of the cmcH genes of Nocardia lactamdurans and Streptomyces clavuligerus encoding a functional 3′-hydroxymethylcephem O-carbamoyltransferase for cephamycin biosynthesis. Gene 162:21–27

    PubMed  Article  CAS  Google Scholar 

  • Coque JJ, Enguita FJ, Cardoza RE, Martin JF, Liras P (1996) Characterization of the cefF gene of Nocardia lactamdurans encoding a 3′-methylcephem hydroxylase different from the 7-cephem hydroxylase. Appl Microbiol Biotechnol 44:605–609

    PubMed  Article  CAS  Google Scholar 

  • Daly JW (2004) Marine toxins and nonmarine toxins: convergence or symbiotic organisms? J Nat Prod 67:1211–1215

    PubMed  Article  CAS  Google Scholar 

  • Du L, Sanchez C, Chen M, Edwards DJ, Shen B (2000) The biosynthetic gene cluster for the antitumor drug bleomycin from Streptomyces verticillus ATCC15003 supporting functional interactions between nonribosomal peptide synthetases and a polyketide synthase. Chem Biol 7:623–642

    PubMed  Article  CAS  Google Scholar 

  • Felsenstein J (1989) PHYLIP. Phylogeny inference package. Cladistics 5:164–166

    Google Scholar 

  • Gallacher S, Flynn KJ, Franco JM, Brueggemann EE, Hines HB (1997) Evidence for production of paralytic shellfish toxins by bacteria associated with Alexandrium spp. (Dinophyta) in culture. Appl Environ Microbiol 63:239–245

    PubMed  CAS  Google Scholar 

  • Gogarten JP, Townsend JP (2005) Horizontal gene transfer, genome innovation and evolution. Nat Rev Microbiol 3:679–687

    PubMed  Article  CAS  Google Scholar 

  • Hackett JD, Scheetz TE, Yoon HS, Soares MB, Bonaldo MF, Casavant TL, Bhattacharya D (2005) Insights into a dinoflagellate genome through expressed sequence tag analysis. BMC Genom 6:80

    Article  Google Scholar 

  • Hallegraeff GM (1995) Harmful algal blooms: A global overview. In: Hallegraeff GM, Anderson DM, Cembella AD (eds) Manual on harmful marine microalgae. UNESCO, Paris, pp 1–22

    Google Scholar 

  • Haydock SF, Appleyard AN, Mironenko T, Lester J, Scott N, Leadlay PF (2005) Organization of the biosynthetic gene cluster for the macrolide concanamycin A in Streptomyces neyagawaensis ATCC 27449. Microbiology 151:3161–3169

    PubMed  Article  CAS  Google Scholar 

  • Henikoff JG, Greene EA, Pietrokovski S, Henikoff S (2000) Increased coverage of protein families with the blocks database servers. Nucleic Acids Res 28:228–230

    PubMed  Article  CAS  Google Scholar 

  • Honda D, Yokota A, Sugiyama J (1999) Detection of seven major evolutionary lineages in cyanobacteria based on the 16S rRNA gene sequence analysis with new sequences of five marine Synechococcus strains. J Mol Evol 48:723–739

    PubMed  Article  CAS  Google Scholar 

  • Jabbouri S, Fellay R, Talmont F, Kamalaprija P, Burger U, Relic B, Prome JC, Broughton WJ (1995) Involvement of nodS in N-methylation and nodU in 6-O-carbamoylation of Rhizobium sp. NGR234 nod factors. J Biol Chem 270:22968–22973

    PubMed  Article  CAS  Google Scholar 

  • Jabbouri S, Relic B, Hanin M, Kamalaprija P, Burger U, Prome D, Prome JC, Broughton WJ (1998) nolO and noeI (HsnIII) of Rhizobium sp. NGR234 are involved in 3-O-carbamoylation and 2-O-methylation of Nod factors. J Biol Chem 273:12047–12055

    PubMed  Article  CAS  Google Scholar 

  • Jukes TH, Cantor CR (1969) Evolution of protein molecules. In: Munro HN (ed) Mammalian protein metabolism. Academic Press, New York, pp 21–132

    Google Scholar 

  • Kaas H, Henriksen P (2000) Saxitoxins (PSP toxins) in Danish lakes. Water Res 34:2089–2097

    Article  CAS  Google Scholar 

  • Kao CY, Levinson SR (1986) Tetrodotoxin, Saxitoxin, and the molecular biology of the sodium channel. In: Boland B, Cullinan J, Cohn T (eds) Annals of the New York Academy of Science. New York Academy of Science, New York, pp 1–445

    Google Scholar 

  • Kellmann R, Neilan BA (2007) Biochemical characterisation of paralytic shellfish toxin biosynthesis in vitro. J Phycol 43:497–508

    Article  CAS  Google Scholar 

  • Kellmann R, Michali TK, Jeon YJ, Pickford R, Pomati F, Neilan BA (2008) Biosynthetic intermediate analysis and functional homolgy reveal a saxitoxin gene cluster in cyanobacteria. Appl Environ Microbiol 74:4044–4053

    PubMed  Article  CAS  Google Scholar 

  • Kharel MK, Basnet DB, Lee HC, Liou K, Woo JS, Kim BG, Sohng JK (2004) Isolation and characterization of the tobramycin biosynthetic gene cluster from Streptomyces tenebrarius. FEMS Microbiol Lett 230:185–190

    PubMed  Article  CAS  Google Scholar 

  • Kodoma M, Ogata T, Sato S (1988) Bacterial production of saxitoxin. Agr Biol Chem 52:1075–1077

    Google Scholar 

  • Kodoma M, Ogata T, Sakamoto S, Sato S, Honda T, Miwatani T (1990) Production of paralytic shellfish toxins by a bacterium Moraxella sp. isolated from Protogonyaulax tamarensis. Toxicon 28:707–714

    Article  Google Scholar 

  • Kotaki Y, Tajiri M, Oshima Y, Yasumoto T (1983) Identification of a calcareous red alga as the primary source of paralytic shellfish toxins in coral reef crabs and gastropods. Bull Jpn Soc Sci Fish [Nippon Suisan Gakkaishi] 49:283–286

    Google Scholar 

  • Lerat E, Daubin V, Ochman H, Moran NA (2005) Evolutionary origins of genomic repertoires in bacteria. PLoS Biol 3:e130

    PubMed  Article  CAS  Google Scholar 

  • Madinabeitia N, Bellogin RA, Buendia-Claveria AM, Camacho M, Cubo T, Espuny MR, Gil-Serrano AM, Lyra MC, Moussaid A, Ollero FJ, Soria-Diaz ME, Vinardell JM, Zeng J, Ruiz-Sainz JE (2002) Sinorhizobium fredii HH103 has a truncated nolO gene due to a -1 frameshift mutation that is conserved among other geographically distant S. fredii strains. Mol Plant Microbe Interact 15:150–159

    PubMed  Article  CAS  Google Scholar 

  • Mao Y, Varoglu M, Sherman DH (1999) Molecular characterization and analysis of the biosynthetic gene cluster for the antitumor antibiotic mitomycin C from Streptomyces lavendulae NRRL 2564. Chem Biol 6:251–263

    PubMed  Article  CAS  Google Scholar 

  • Moffitt MC, Neilan BA (2004) Characterization of the nodularin synthetase gene cluster and proposed theory of the evolution of cyanobacterial hepatotoxins. Appl Environ Microbiol 70:6353–6362

    PubMed  Article  CAS  Google Scholar 

  • Nakashima K, Arakawa O, Taniyama S, Nonaka M, Takatani T, Yamamori K, Fuchi Y, Noguchi T (2004) Occurrence of saxitoxins as a major toxin in the ovary of a marine puffer Arothron firmamentum. Toxicon 43:207–212

    PubMed  Article  CAS  Google Scholar 

  • Neilan BA (1995) Identification and phylogenetic analysis of toxigenic cyanobacteria by multiplex randomly amplified polymorphic DNA PCR. Appl Environ Microbiol 61:2286–2291

    PubMed  CAS  Google Scholar 

  • Neilan BA, Jacobs D, DelDot T, Blackall LL, Hawkins PR, Cox PT, Goodman AE (1997) rRNA sequences and evolutionary relationships among toxic and nontoxic cyanobacteria of the genus Microcystis. Int J Syst Bacteriol 47:693–697

    PubMed  CAS  Article  Google Scholar 

  • Onodera H, Satake M, Oshima Y, Yasumoto T, Carmichael Wayne W (1997) New saxitoxin analogues from the freshwater filamentous cyanobacterium Lyngbya wollei. Nat Toxins 5:146–151

    PubMed  CAS  Article  Google Scholar 

  • Oshima Y, Kotaki Y, Harada T, Yasumoto T (1984) Paralytic shellfish toxins in tropical waters. In: Ragelis E (ed) Seafood toxins. American Chemical Society, Washington, DC, pp 160–170

    Google Scholar 

  • Pereira P, Onodera H, Andrinolo D, Franca S, Araujo F, Lagos N, Oshima Y (2000) Paralytic shellfish toxins in the freshwater cyanobacterium Aphanizomenon flos-aquae, isolated from Montargil reservoir, Portugal. Toxicon 38:1689–1702

    PubMed  Article  CAS  Google Scholar 

  • Perrière G, Gouy M (1996) WWW-Query: an on-line retrieval system for biological sequence banks. Biochemie 78:364–369

    Article  Google Scholar 

  • Plumley FG (2001) Purification of an enzyme involved in saxitoxin synthesis. J Phycol 37:926–928

    Article  CAS  Google Scholar 

  • Pomati F, Neilan BA (2004) PCR-based positive hybridization to detect genomic diversity associated with bacterial secondary metabolism. Nucleic Acids Res 32:e7

    PubMed  Article  Google Scholar 

  • Pomati F, Burns BP, Neilan BA (2004) Identification of an Na(+)-dependent transporter associated with saxitoxin-producing strains of the cyanobacterium Anabaena circinalis. Appl Environ Microbiol 70:4711–4719

    PubMed  Article  CAS  Google Scholar 

  • Pomati F, Kellmann R, Burns BP, Cavaliere R, Neilan BA (2006) Comparative gene expression studies of PSP-toxins producing and non-toxic Anabaena circinalis strains and effects of lidocaine hydrochloride. Environ Int 32:734–748

    Article  Google Scholar 

  • Rascher A, Hu Z, Buchanan GO, Reid R, Hutchinson CR (2005) Insights into the biosynthesis of the benzoquinone ansamycins geldanamycin and herbimycin, obtained by gene sequencing and disruption. Appl Environ Microbiol 71:4862–4871

    PubMed  Article  CAS  Google Scholar 

  • Rice P, Longden I, Bleasby A (2000) EMBOSS: The European Molecular Biology Open Software Suite. Trends Genet 16:276–277

    PubMed  Article  CAS  Google Scholar 

  • Saitou N, Nei M (1987) The neighbour-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425

    PubMed  CAS  Google Scholar 

  • Sato S, Kodoma M, Ogata T, Saitanu K, Furuya M, Hirayama K, Kakinuma K (1997) Saxitoxin as a toxic principle of a freshwater puffer Tetraodon fugi in Thailand. Toxicon 35:137–140

    PubMed  Article  CAS  Google Scholar 

  • Shimizu Y (1993) Microalgal metabolites. Chem Rev 93:1685–1698

    Article  CAS  Google Scholar 

  • Siebert PD, Chenchik A, Kellogg DE, Lukyanov KA, Lukyanov SA (1995) An improved PCR method for walking in uncloned genomic DNA. Nucleic Acids Res 23:1087–1088

    PubMed  Article  CAS  Google Scholar 

  • Sivonen K (1996) Cyanobacterial toxins and toxin production. Phycologia 35:12–24

    Google Scholar 

  • Steffensky M, Muhlenweg A, Wang ZX, Li SM, Heide L (2000) Identification of the novobiocin biosynthetic gene cluster of Streptomyces spheroides NCIB 11891. Antimicrob Agents Chemother 44:1214–1222

    PubMed  Article  CAS  Google Scholar 

  • Su Z, Sheets M, Ishida H, Li F, Barry WH (2004) Saxitoxin blocks L-type ICa. J Pharmacol Exp Ther 308(1):324–329

    PubMed  Article  CAS  Google Scholar 

  • Thompson AS, Rhodes JC, Pettman I (1988) Catalogue of strains. Natural Environment Research Council Culture Collection of Algae and Protozoa, p 22

  • Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 25:4876–4882

    PubMed  Article  CAS  Google Scholar 

  • Wang J, Salata JJ, Bennett PB (2003) Saxitoxin is a gating modifier of HERG K+ channels. J Gen Physiol 121:583–598

    PubMed  Article  CAS  Google Scholar 

  • Yotsu-Yamashita M, Kim YH, Dudley SC Jr, Choudhary G, Pfahnl A, Oshima Y, Daly JW (2004) The structure of zetekitoxin AB, a saxitoxin analog from the Panamanian golden frog Atelopus zeteki. Proc Natl Acad Sci USA 101:4346–4351

    PubMed  Article  CAS  Google Scholar 

  • Yu TW, Bai L, Clade D, Hoffmann D, Toelzer S, Trinh KQ, Xu J, Moss SJ, Leistner E, Floss HG (2002) The biosynthetic gene cluster of the maytansinoid antitumor agent ansamitocin from Actinosynnema pretiosum. Proc Natl Acad Sci USA 99:7968–7973

    PubMed  Article  CAS  Google Scholar 

  • Zaman L, Arakawa O, Shimosu A, Onoue Y (1997) Occurrence of paralytic shellfish poison in Bangladeshi freshwater puffers. Toxicon 35:423–431

    PubMed  Article  CAS  Google Scholar 

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Acknowledgments

The Australian Research Council is thanked for its financial support. Wayne Carmichael, Peter Baker, and Martin Saker are thanked for providing cyanobacterial samples.

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Affiliations

  1. Department of Molecular Biology, University of Bergen, P.O. Box 7803, 5020, Bergen, Norway

    Ralf Kellmann

  2. School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, NSW, 205, Australia

    Troco Kaan Michali & Brett Adam Neilan

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  1. Ralf Kellmann
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  2. Troco Kaan Michali
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Correspondence to Ralf Kellmann.

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An erratum to this article can be found at http://dx.doi.org/10.1007/s00239-009-9210-0

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Kellmann, R., Michali, T.K. & Neilan, B.A. Identification of a Saxitoxin Biosynthesis Gene with a History of Frequent Horizontal Gene Transfers. J Mol Evol 67, 526–538 (2008). https://doi.org/10.1007/s00239-008-9169-2

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  • DOI: https://doi.org/10.1007/s00239-008-9169-2

Keywords

  • Paralytic shellfish poisoning
  • Saxitoxin
  • Biosynthesis gene
  • O-Carbamoyltransferase
  • Horizontal gene transfer
  • Phylogeny