Antitumor polyketide biosynthesis by an uncultivated bacterial symbiont of the marine sponge Theonella swinhoei

Abstract

Bacterial symbionts have long been suspected to be the true producers of many drug candidates isolated from marine invertebrates. Sponges, the most important marine source of biologically active natural products, have been frequently hypothesized to contain compounds of bacterial origin. This symbiont hypothesis, however, remained unproven because of a general inability to cultivate the suspected producers. However, we have recently identified an uncultured Pseudomonas sp. symbiont as the most likely producer of the defensive antitumor polyketide pederin in Paederus fuscipes beetles by cloning the putative biosynthesis genes. Here we report closely related genes isolated from the highly complex metagenome of the marine sponge Theonella swinhoei, which is the source of the onnamides and theopederins, a group of polyketides that structurally resemble pederin. Sequence features of the isolated genes clearly indicate that it belongs to a prokaryotic genome and should be responsible for the biosynthesis of almost the entire portion of the polyketide structure that is correlated with antitumor activity. Besides providing further proof for the role of the related beetle symbiont-derived genes, these findings raise intriguing ecological and evolutionary questions and have important general implications for the sustainable production of otherwise inaccessible marine drugs by using biotechnological strategies.

Fig. 1.

Some members of the pederin family of antitumor compounds. 1, Pederin from Paederus spp. rove beetles; 2, theopederin A from sponge T. swinhoei; 3, onnamide A from T. swinhoei.

Fig. 2.

Comparison of the onn gene cluster with the ped system and proposed onnamide A biosynthesis. (A) Map of the ped cluster from the Pa. fuscipes symbiont. Double slashes separate the three genome regions. (B) Map of the onn cluster and correlation to ped homologs. (C) Protein products of the PKS-NRPS genes onnB and onnI and proposed biosynthetic pathway leading to onnamide/theopederin-type compounds. Each circle represents one domain. The intermediates are attached to ACP or, in the case of the second module of OnnI, peptidyl carrier protein (PCP) domains (shown as small, filled circles). Domains in gray lack active site motifs and are presumably nonfunctional. CR, CR superfamily; EST, esterase; REG, regulator; OXY, oxygenase; OR, oxidoreductase; TP, transposase; C, condensation domain; A, adenylation domain.

Fig. 3.

PCR analysis of PKS genes in different specimens of T. swinhoei. B, blind control without template DNA; W2, T. swinhoei W2 (onnamide-free); Y3, T. swinhoei Y3 (onnamide-free); Y1 and Y4, two different onnamide-containing specimens of T. swinhoei Y; pks3, TSY1-3-specific PKS primers sponge3f/sponge3r; pks11, TSY1-11-specific PKS primers sponge11f/sponge11r.