Synteruptor: mining genomic islands for non-classical specialized metabolite gene clusters

Abstract

Microbial specialized metabolite biosynthetic gene clusters (SMBGCs) are a formidable source of natural products of pharmaceutical interest. With the multiplication of genomic data available, very efficient bioinformatic tools for automatic SMBGC detection have been developed. Nevertheless, most of these tools identify SMBGCs based on sequence similarity with enzymes typically involved in specialised metabolism and thus may miss SMBGCs coding for undercharacterised enzymes. Here we present Synteruptor (https://bioi2.i2bc.paris-saclay.fr/synteruptor), a program that identifies genomic islands, known to be enriched in SMBGCs, in the genomes of closely related species. With this tool, we identified a SMBGC in the genome of Streptomyces ambofaciens ATCC23877, undetected by antiSMASH versions prior to antiSMASH 5, and experimentally demonstrated that it directs the biosynthesis of two metabolites, one of which was identified as sphydrofuran. Synteruptor is also a valuable resource for the delineation of individual SMBGCs within antiSMASH regions that may encompass multiple clusters, and for refining the boundaries of these SMBGCs.

Figure 1.

Outline of the pipeline used for the identification of the genomic islands. From annotated genomes, ortholog pairs are determined. Synteny blocks are constructed by clustering all consecutive orthologs and synteny breaks (genomic islands) between two consecutive synteny blocks are identified. The genomic islands are then analysed to determine a number of parameters such as number of CDSs, number of CDSs without ortholog. BRH is for Best reciprocal BLAST hits.

Figure 2.

Examples of the output visualization graphs and tables obtained when comparing the genomes of S. ambofaciens ATCC 23877 and S. coelicolor A(3)2. Only genomic islands of at least 15 CDSs found in at least one genome are shown. (A) Dotplot showing ortholog pairs and synteny blocks, with the genomic islands circled in red. GOC (Gene Order Conservation) profiles are displayed below or on the left of the dotplot. The scale on the x-axis indicates the gene position in gene number, not in bp. (B) Close-up of the central genomic regions. The red boxes represent the GIs observed either in the genome of S. coelicolor (vertical bars) or in the genome of S. ambofaciens (horizontal bars) The arrow indicates the genomic island visualized in (C). (C) Visualization of the gene content and organization of the genomic island 0762fd (#819). Genes in dark colours do not possess orthologs in the other genome, genes in light colours do. Green bars represent tRNA genes. Left and right blocks represent synteny regions. This figure represents snapshots of the web interface. (D) Tables describing the genes located in the left synteny blocks of the 0762fd genomic island. The gene ID, the position in the chromosome, the difference of the GC ratio between the gene and the complete genome sequence, as well as the predicted function and length of the gene product are presented. In this example, two genes, one in each genome (SAM23877_3917 and SCO3725), have a paralog in their respective chromosome. Num: CDS number. These tables represent a snapshot of the web interface. (E) Number of genomic islands detected in S. ambofaciens ATCC23877 genome when compared to S. coelicolor A3(2) related to the minimal number of CDSs in the genomic islands.

Figure 3.

(A) HPLC analysis of culture supernatants of S. ambofaciens ATCC 23877 OSC4 and OSC416 (SAM23877_3931 inactivated), S. coelicolor M1154 and S. coelicolor SPFSH001. First column, ELSD monitoring; second column UV monitoring at 245 nm. (B) Scheme of the chemical degradation of sphydrofuran (1) into 2-methyl-4-(1-glycerol)-furan (2)

Figure 4.

Example of a hotspot of integration detected by Synteruptor. (A) Gene content and organization of the genomic islands in the synteny break 478e7e (#807); (B) break graph representing the existence of genomic islands (link between two organisms) located at the same position in the genomes of the database as the genomic islands observed in (A); and (C) Visualization of the genomic islands located at the same position than the genomic islands observed in (A). (D) Illustration of overlapping breaks for the genomic island presented in (A), showing that in S. lunaelactis MM109 and Streptomyces sp. CC0208 genomes, genomic islands are present in the same chromosomal region, but the boundaries of these islands are slightly different.

Figure 5.

Localization of S. ambofaciens ATCC 23877 genomic islands and SMBGCs; A. Screenshot of Synteruptor showing the dotplot and GOC profile when comparing S. ambofaciens ATCC23877 and S. coelicolor A(3)2 chromosomes and looking for genomic islands of three or more CDS in S. ambofaciens. Dotted lines indicate the approximate limits of the synteny region with the chromosome of S. coelicolor A(3)2.; B. Schematic representation of the S. ambofaciens ATCC23877 chromosome with the location of the known SMBGCs and of the newly identified one, sphydrofuran. The arms are represented by boxes filled with a blue gradient.