Figure 1.
Secondary metabolic pathways from three Prochloron metagenomes.
Center: Prochloron cells (top) from the ascidian L. patella (bottom). Surrounding are ten pathway types identified, comprising 15 distinct biosynthetic gene clusters in the metagenomes of P1, P2 and P3. Compounds that have been chemically isolated are indicated by an asterisk; the remainder are genome-based predictions.
Figure 2.
Variations in secondary metabolic gene clusters constitute the major difference between the P. didemni genomes.
a) The Artemis Comparison Tool (ACT) was used to visualize pair-wise comparisons between contig ASM-2318 from P1 and corresponding contigs from P2 and P3 [44]. Violet color indicates ≥ 96% nucleotide identity while light blue color indicates 90–96% nucleotide identity. Two of the three differing regions marked by black boxes differ solely due to natural product gene clusters. This pattern is reproduced throughout the 3 metagenomes. b) and c) Dot Plot representations of NUCmer comparisons showing synteny between the same genetic regions as in A [45].
Figure 3.
Schematic representation of the different levels of changes in functional cassettes in P. didemni genomes.
Three levels can be observed: 1. Presence or absence of a gene cluster in the genome. Homologous gene clusters are >99% identical between strains; 2. Swaps of function (heterocyclization to prenylation) within otherwise >99% identical gene clusters; 3. Functional point mutations in gene clusters (for example in product-coding sequences of ribosomal peptides).
Figure 4.
Sporadic distribution of ascidian-associated secondary metabolic genes across the tropical Pacific.
Twenty-four samples collected over a 5,500 km transect in the tropical Pacific were screened for presence/absence of genes for five biosynthetic pathways. Each circle represents an individual ascidian sample. The shape inside the circle indicates the species of the Prochloron-harboring ascidian, as shown in the key above. Colors within the circle represent the presence of individual biosynthetic pathways, which are >99% identical between samples, while black indicates the absence of a pathway from the sample. S represents samples for which whole genome sequence was obtained.
Figure 5.
New prenylated cyanobactins discovered by comparative analysis.
A control known compound, trunkamide (3, panel a), was subjected to high-resolution mass spectrometry analysis. Using tandem mass spectrometry, both one and two isoprene side chains (C5H8, m/z = 68.0626) were eliminated from this molecule. In comparison, two new compounds, mollamide D (1) and mollamide E (2) from sample 03-002, also exhibited the same pattern (panels b and c). The high-resolution results corresponded both to the predicted masses of the compounds and to the predicted loss of isoprene.