Variable genetic architectures produce virtually identical molecules in bacterial symbionts of fungus-growing ants

Abstract

Small molecules produced by Actinobacteria have played a prominent role in both drug discovery and organic chemistry. As part of a larger study of the actinobacterial symbionts of fungus-growing ants, we discovered a small family of three previously unreported piperazic acid-containing cyclic depsipeptides, gerumycins A-C. The gerumycins are slightly smaller versions of dentigerumycin, a cyclic depsipeptide that selectively inhibits a common fungal pathogen, Escovopsis. We had previously identified this molecule from a Pseudonocardia associated with Apterostigma dentigerum, and now we report the molecule from an associate of the more highly derived ant Trachymyrmex cornetzi. The three previously unidentified compounds, gerumycins A-C, have essentially identical structures and were produced by two different symbiotic Pseudonocardia spp. from ants in the genus Apterostigma found in both Panama and Costa Rica. To understand the similarities and differences in the biosynthetic pathways that produced these closely related molecules, the genomes of the three producing Pseudonocardia were sequenced and the biosynthetic gene clusters identified. This analysis revealed that dramatically different biosynthetic architectures, including genomic islands, a plasmid, and the use of spatially separated genetic loci, can lead to molecules with virtually identical core structures. A plausible evolutionary model that unifies these disparate architectures is presented.

Keywords: biosynthetic gene clusters; chemical ecology; horizontal gene transfer; natural products; symbiosis.

Fig. 1.

Cyclic depsipeptides from ant-associated Pseudonocardia spp. The atoms highlighted in red in molecules 2 and 3 are derived from a spatially separated BGC.

Fig. 2.

BGCs encoding the production of 1–4. The hybrid NRPS/PKS gene cluster responsible for the production of 1 is drawn, highlighting the juxtaposition of different subclusters (NRPS in green; PKS in blue). The two distinct plasmid-borne BGCs (named primary and accessory) are separated by ∼92 kb in EC080625-04 and encode for the full suite of enzymes required to produce 2 and 3. The single chromosomal BGC in HH130629-09 that encodes for 4 is drawn, highlighting the primary and accessory plasmid-borne clusters.

Fig. 3.

Plausible scheme of BGC evolution in an ant-associated bacterium. We depict the stepwise unification of plasmid-borne subclusters and integration into the bacterial chromosome. The scheme is representative of the transition from a plasmid-borne split BGC in EC080625-04 to a newly integrated singular version of these BGCs in HH130629-09. A hypothetical plasmid-borne intermediate (plasmid*) is also drawn.