title: Novel family of terpene synthases evolved from trans-isoprenyl diphosphate synthases in a flea beetle.
abstract: Sesquiterpenes play important roles in insect communication, for example as pheromones. However, no sesquiterpene synthases, the enzymes involved in construction of the basic carbon skeleton, have been identified in insects to date. We investigated the biosynthesis of the sesquiterpene (6R,7S)-himachala-9,11-diene in the crucifer flea beetle Phyllotreta striolata, a compound previously identified as a male-produced aggregation pheromone in several Phyllotreta species. A (6R,7S)-himachala-9,11-diene-producing sesquiterpene synthase activity was detected in crude beetle protein extracts, but only when (Z,E)-farnesyl diphosphate [(Z,E)-FPP] was offered as a substrate. No sequences resembling sesquiterpene synthases from plants, fungi, or bacteria were found in the P. striolata transcriptome, but we identified nine divergent putative trans-isoprenyl diphosphate synthase (trans-IDS) transcripts. Four of these putative trans-IDSs exhibited terpene synthase (TPS) activity when heterologously expressed. Recombinant PsTPS1 converted (Z,E)-FPP to (6R,7S)-himachala-9,11-diene and other sesquiterpenes observed in beetle extracts. RNAi-mediated knockdown of PsTPS1 mRNA in P. striolata males led to reduced emission of aggregation pheromone, confirming a significant role of PsTPS1 in pheromone biosynthesis. Two expressed enzymes showed genuine IDS activity, with PsIDS1 synthesizing (E,E)-FPP, whereas PsIDS3 produced neryl diphosphate, (Z,Z)-FPP, and (Z,E)-FPP. In a phylogenetic analysis, the PsTPS enzymes and PsIDS3 were clearly separated from a clade of known coleopteran trans-IDS enzymes including PsIDS1 and PsIDS2. However, the exon-intron structures of IDS and TPS genes in P. striolata are conserved, suggesting that this TPS gene family evolved from trans-IDS ancestors.
authors.affiliation: Research Group Sequestration and Detoxification in Insects, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany; fberan@ice.mpg.de.
authors.forename: Franziska
authors.initials: F
authors.lastname: Beran
authors.affiliation: Department of Bioorganic Chemistry, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany;
authors.forename: Peter
authors.initials: P
authors.lastname: Rahfeld
authors.affiliation: Department of Biochemistry, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany;
authors.forename: Katrin
authors.initials: K
authors.lastname: Luck
authors.affiliation: Department of Biochemistry, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany;
authors.forename: Raimund
authors.initials: R
authors.lastname: Nagel
authors.affiliation: Department of Entomology, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany;
authors.forename: Heiko
authors.initials: H
authors.lastname: Vogel
authors.affiliation: Department of Mass Spectrometry, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany;
authors.forename: Natalie
authors.initials: N
authors.lastname: Wielsch
authors.affiliation: Department of Biochemistry, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany;
authors.forename: Sandra
authors.initials: S
authors.lastname: Irmisch
authors.affiliation: Entomology, AVRDC-The World Vegetable Center, Shanhua, Tainan 74151, Taiwan.
authors.forename: Srinivasan
authors.initials: S
authors.lastname: Ramasamy
authors.affiliation: Department of Biochemistry, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany;
authors.forename: Jonathan
authors.initials: J
authors.lastname: Gershenzon
authors.affiliation: Department of Entomology, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany;
authors.forename: David G
authors.initials: DG
authors.lastname: Heckel
authors.affiliation: Department of Biochemistry, Max Planck Institute for Chemical Ecology, 07745 Jena, Germany;
authors.forename: Tobias G
authors.initials: TG
authors.lastname: Köllner
mesh_terms: D019883:Alkyl and Aryl Transferases / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D001517:Coleoptera / Q000145:classification / Q000201:enzymology* / Q000235:genetics; D019143:Evolution, Molecular; D005260:Female; D040461:Gene Components; D017344:Genes, Insect; D049810:Genetic Speciation; D019476:Insect Proteins / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D008297:Male; D008969:Molecular Sequence Data; D005810:Multigene Family; D016366:Open Reading Frames / Q000235:genetics; D010675:Pheromones / Q000096:biosynthesis*; D010802:Phylogeny; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012717:Sesquiterpenes / Q000378:metabolism*; D059467:Transcriptome
chemical_list: C000609685:(6R,7S)-himachala-9,11-diene; D019476:Insect Proteins; D010675:Pheromones; D011993:Recombinant Fusion Proteins; D012717:Sesquiterpenes; D019883:Alkyl and Aryl Transferases; C108877:terpene synthase
keywords: (6R,7S)-himachala-9,11-diene; aggregation pheromone; flea beetle; isoprenyl diphosphate synthase; sesquiterpene synthase