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. 2009 Jun 1;4(6):e5758.
doi: 10.1371/journal.pone.0005758.

Origin of Saxitoxin Biosynthetic Genes in Cyanobacteria

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Free PMC article

Origin of Saxitoxin Biosynthetic Genes in Cyanobacteria

Ahmed Moustafa et al. PLoS One. .
Free PMC article

Abstract

Background: Paralytic shellfish poisoning (PSP) is a potentially fatal syndrome associated with the consumption of shellfish that have accumulated saxitoxin (STX). STX is produced by microscopic marine dinoflagellate algae. Little is known about the origin and spread of saxitoxin genes in these under-studied eukaryotes. Fortuitously, some freshwater cyanobacteria also produce STX, providing an ideal model for studying its biosynthesis. Here we focus on saxitoxin-producing cyanobacteria and their non-toxic sisters to elucidate the origin of genes involved in the putative STX biosynthetic pathway.

Methodology/principal findings: We generated a draft genome assembly of the saxitoxin-producing (STX+) cyanobacterium Anabaena circinalis ACBU02 and searched for 26 candidate saxitoxin-genes (named sxtA to sxtZ) that were recently identified in the toxic strain Cylindrospermopsis raciborskii T3. We also generated a draft assembly of the non-toxic (STX-) sister Anabaena circinalis ACFR02 to aid the identification of saxitoxin-specific genes. Comparative phylogenomic analyses revealed that nine putative STX genes were horizontally transferred from non-cyanobacterial sources, whereas one key gene (sxtA) originated in STX+ cyanobacteria via two independent horizontal transfers followed by fusion. In total, of the 26 candidate saxitoxin-genes, 13 are of cyanobacterial provenance and are monophyletic among the STX+ taxa, four are shared amongst STX+ and STX-cyanobacteria, and the remaining nine genes are specific to STX+ cyanobacteria.

Conclusions/significance: Our results provide evidence that the assembly of STX genes in ACBU02 involved multiple HGT events from different sources followed presumably by coordination of the expression of foreign and native genes in the common ancestor of STX+ cyanobacteria. The ability to produce saxitoxin was subsequently lost multiple independent times resulting in a nested relationship of STX+ and STX- strains among Anabaena circinalis strains.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Phylogenetic tree of 16S rRNA from cyanobacteria.
Unrooted ML phylogenetic trees inferred from small subunit (16S) rRNA that include (a) different cyanobacterial orders and (b) only saxitoxin-producing (STX+) and STX− strains of Anabaena circinalis. ML bootstrap values (when ≥50%) are indicated at the nodes. The branch lengths are proportional to the number of substitutions per site (see scales in the figure). The toxic and non-toxic strains are indicated by the plus sign (+) and minus sign (−), respectively.
Figure 2
Figure 2. Phylogeny of sxtY and sxtZ.
ML phylogenetic trees of (a) sxtY (phosphate uptake regulator) and (b) sxtZ (histidine kinase). These represent the class of saxitoxin-related genes of cyanobacterial origin in which taxa most closely related to the STX+ strains are members of the Nostocaceae. ML bootstrap values (when ≥50%) are indicated at the nodes. The branch lengths are proportional to the number of substitutions per site (see scales in the figure). Cyanobacterial taxa are shown in blue text, non-cyanobacterial prokaryotes in black text, and eukaryotes (Plantae) in green except for the photosynthetic filose amoeba Paulinella chromatophora in red. The trees have been rooted arbitrarily.
Figure 3
Figure 3. Phylogeny of sxtN.
ML phylogenetic tree of sxtN (sulfotransferase) that represents saxitoxin-related genes of cyanobacterial origin that is potentially from members of the Oscillatoriales. ML bootstrap values (when ≥50%) are indicated at the nodes. The branch lengths are proportional to the number of substitutions per site (see scale in the figure). Cyanobacterial taxa are shown in blue text, non-cyanobacterial prokaryotes in black text, and eukaryotes (Plantae) in green. The tree has been rooted on the branch leading to the non-cyanobacterial prokaryotes.
Figure 4
Figure 4. Gene fusion origin of sxtA.
(a) Schematic diagram illustrating the fusion event that resulted in sxtA (polyketide synthase) in STX+ cyanobacteria. (b) ML phylogenetic tree of sxtA after dividing it into its two phylogenetic components; i.e., sxtA1 (a gene encoding acyl-CoA N-acyltransferase and phosphopantetheine binding domain-containing protein) and sxtA2 (a gene encoding aminotransferase class I and II). ML bootstrap values (when ≥50%) are indicated at the nodes. The branch lengths are proportional to the number of substitutions per site (see scale in the figure). Cyanobacterial taxa are shown in blue text, actinobacteria in brown text, proteobacteria in red text, and other non-cyanobacterial prokaryotes in black. The tree was rooted on the branch separating the two gene fusion partners.
Figure 5
Figure 5. Origin of ACBU02 genes.
Venn diagram showing the genome-wide distribution of genes in the STX+ cyanobacterium Anabaena circinalis ACBU02 with respect to sequences shared with other prokaryotes and viruses.
Figure 6
Figure 6. Phylogeny of SAM-dependent methyltransferase.
Unrooted ML phylogenetic tree of a putative STX+ specific S-adenosyl-L-methionine (SAM)-dependent methyltransferase that has a potential Bacillus-like origin. ML bootstrap values (when ≥50%) are indicated at the nodes. The branch lengths are proportional to the number of substitutions per site (see scale in the figure). Cyanobacterial taxa are shown in blue text, non-cyanobacterial prokaryotes in black text, and eukaryotes (Plantae) in green.

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