Characterization of a bark beetle oxidoreductase with pheromone-biosynthetic expression patterns
Rubi D. Figueroa Teran, email@example.com, Gary J. Blomquist, William Welch, and Claus Tittiger. University of Nevada, Reno, Department of Biochemistry and Molecular Biology, MS330, Reno, NV
Enantiomeric blends of ipsdienol (2-methyl-6-methylene-2,7-octadien-4-ol) and ipsenol (2-methyl-6-methylene-7-octen-4-ol) are the major pheromones of Ips species (Coleoptera: Scolytidae). They are biosynthesized through the mevalonate pathway to myrcene, which is then hydroxylated to ipsdienol. The last steps of pheromone biosynthesis are unknown, but are thought to be catalyzed by oxidoreductases and include the oxidation of ipsdienol to ipsdienone followed by stereo-specific reduction to ipsdienol. Microarray and quantitative real-time PCR analyses identified an oxidoreductase, represented by IPG012D04 in our EST library of the western pine engraver beetles, (Ipspini), which exhibits pheromone-biosynthetic expression patterns. These include high basal levels in adult males compared to fed and juvenile hormone-treated females and highly upregulated expression in the anterior midgut, the pheromone producing tissue of fed males. Full-length cDNAs of this oxidoreductase and homologues from eastern I.pini and the pinyon ips (Ipsconfusus) have been cloned and sequenced. The western I.pini cDNA encoding the enzyme was cloned into the BaculoDirect Baculovirus vector for protein expression in Sf9 cells. An NADP(H) binding assay is being optimized to characterize the function of the oxidoreductase. Deuterated substrates will be used for further functional analysis of formed products using coupled GC-MS. Comparing the products formed by each homolog will determine the final steps of pheromone biosynthesis and provide insight into how Ipsspp. produce different enantiomeric blends of the same compounds.
Species 1: Coleoptera Curculionidae Ipspini (pine engraver) Species 2: Coleoptera Curculionidae Ipsconfusus (pinyon ips)