A novel fatty acyl desaturase from the pheromone glands of Ctenopseustis obliquana and C. herana with specific Z5-desaturase activity on myristic acid

Abstract

Sexual communication in the Lepidoptera typically involves a female-produced sex pheromone that attracts males of the same species. The most common type of moth sex pheromone comprises individual or blends of fatty acyl derivatives that are synthesized by a specific enzymatic pathway in the female's pheromone gland, often including a desaturation step. This reaction is catalyzed by fatty acyl desaturases that introduce double bonds at specific locations in the fatty acid precursor backbone. The two tortricid moths, Ctenopseustis obliquana and C. herana (brown-headed leafrollers), which are endemic in New Zealand, both use (Z)-5-tetradecenyl acetate as part of their sex pheromone. In C. herana, (Z)-5-tetradecenyl acetate is the sole component of the pheromone. Labeling experiments have revealed that this compound is produced via an unusual Δ5-desaturation of myristic acid. Previously six desaturases were identified from the pheromone glands of Ctenopseustis and its sibling genus Planotortrix, with one differentially regulated to produce the distinct blends used by individual species. However, none were able to conduct the Δ5-desaturation observed in C. herana, and presumably C. obliquana. We have now identified an additional desaturase gene, desat7, expressed in the pheromone glands of both Ctenopseustis species, which is not closely related to any previously described moth pheromone desaturase. The encoded enzyme displays Δ5-desaturase activity on myristic acid when heterologously expressed in yeast, but is not able to desaturate any other fatty acid (C8-C16). We conclude that desat7 represents a new group of desaturases that has evolved a role in the biosynthesis of sex pheromones in moths.

Fig. 1

Sequence and phylogenetic analysis of Desat7. a An alignment of Desat7 isoforms from Ctenopseustis obliquana and C. herana. Amino acid differences are displayed in white or light grey. b A phylogenetic tree including Desat7 (marked with black triangles) together with other moth desaturases as well as desaturases from other organisms (SI Table 1). Groups containing desaturases with similar activities are indicated with brackets and all bootstrap values are noted at the node of each branch

Fig. 2

Quantitative RT-PCR of desat7 in pheromone gland and abdominal tissues of Ctenopseustis obliquana and C. herana adult females. Co Ctenopseustis obliquana, Ch C. herana, PG pheromone gland, Ab abdominal tissue, BLD below limits of detection. Error bars are standard errors of the means of three biological replicates

Fig. 3

GC/MS Analyses of products from heterologous expression of Desat7 in Saccharomyces cerevisia.e a The top panel corresponds to the base methanolysed extract of yeast expressing Desat7 from Ctenopseustis obliquana, allele 38. The enzyme product (Z)-5-tetradecenoate methyl ester (Z5-14:ME) is indicated. The bottom panel shows the extract from the negative control with no Z5-14:ME produced. b The reference compounds Z5-14:ME (top panel) and (E)-5-tetradecenyl methyl ester (E5-14:ME) (bottom panel). The difference in retention time is indicated with dashed lines. c DMDS adducts of the extract from a, with the extracted ions 334, 173, and 161 indicative of the Δ5 double bond (top panel) that is not present in the negative control (bottom panel)