title: Meta-omic characterization of the marine invertebrate microbial consortium that produces the chemotherapeutic natural product ET-743.
abstract: In many macroorganisms, the ultimate source of potent biologically active natural products has remained elusive due to an inability to identify and culture the producing symbiotic microorganisms. As a model system for developing a meta-omic approach to identify and characterize natural product pathways from invertebrate-derived microbial consortia, we chose to investigate the ET-743 (Yondelis) biosynthetic pathway. This molecule is an approved anticancer agent obtained in low abundance (10(-4)-10(-5) % w/w) from the tunicate Ecteinascidia turbinata and is generated in suitable quantities for clinical use by a lengthy semisynthetic process. On the basis of structural similarities to three bacterial secondary metabolites, we hypothesized that ET-743 is the product of a marine bacterial symbiont. Using metagenomic sequencing of total DNA from the tunicate/microbial consortium, we targeted and assembled a 35 kb contig containing 25 genes that comprise the core of the NRPS biosynthetic pathway for this valuable anticancer agent. Rigorous sequence analysis based on codon usage of two large unlinked contigs suggests that Candidatus Endoecteinascidia frumentensis produces the ET-743 metabolite. Subsequent metaproteomic analysis confirmed expression of three key biosynthetic proteins. Moreover, the predicted activity of an enzyme for assembly of the tetrahydroisoquinoline core of ET-743 was verified in vitro. This work provides a foundation for direct production of the drug and new analogues through metabolic engineering. We expect that the interdisciplinary approach described is applicable to diverse host-symbiont systems that generate valuable natural products for drug discovery and development.
authors.affiliation: Life Sciences Institute, University of Michigan, Ann Arbor, Michigan 48109, United States. rmw@lamar.colostate.edu
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Rath
authors.forename: Benjamin
authors.initials: B
authors.lastname: Janto
authors.forename: Josh
authors.initials: J
authors.lastname: Earl
authors.forename: Azad
authors.initials: A
authors.lastname: Ahmed
authors.forename: Fen Z
authors.initials: FZ
authors.lastname: Hu
authors.forename: Luisa
authors.initials: L
authors.lastname: Hiller
authors.forename: Meg
authors.initials: M
authors.lastname: Dahlgren
authors.forename: Rachael
authors.initials: R
authors.lastname: Kreft
authors.forename: Fengan
authors.initials: F
authors.lastname: Yu
authors.forename: Jeremy J
authors.initials: JJ
authors.lastname: Wolff
authors.forename: Hye Kyong
authors.initials: HK
authors.lastname: Kweon
authors.forename: Michael A
authors.initials: MA
authors.lastname: Christiansen
authors.forename: Kristina
authors.initials: K
authors.lastname: Håkansson
authors.forename: Robert M
authors.initials: RM
authors.lastname: Williams
authors.forename: Garth D
authors.initials: GD
authors.lastname: Ehrlich
authors.forename: David H
authors.initials: DH
authors.lastname: Sherman
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000737:chemistry / Q000378:metabolism*; D001688:Biological Products / Q000737:chemistry / Q000378:metabolism*; D004149:Dioxoles / Q000737:chemistry / Q000378:metabolism*; D015723:Gene Library; D054892:Metagenome; D059013:Microbial Consortia / Q000235:genetics / Q000502:physiology*; D008968:Molecular Conformation; D010802:Phylogeny; D040901:Proteomics; D017422:Sequence Analysis, DNA; D044005:Tetrahydroisoquinolines / Q000737:chemistry / Q000378:metabolism*; D000077606:Trabectedin; D014561:Urochordata / Q000235:genetics / Q000382:microbiology*
chemical_list: D000970:Antineoplastic Agents; D001688:Biological Products; D004149:Dioxoles; D044005:Tetrahydroisoquinolines; D000077606:Trabectedin