RESUMO
We combine the use of labeled precursors with enzyme inhibitors to decipher the biosynthetic pathway of pheromone biosynthesis and the rate-limiting step/s that are regulated by pheromone biosynthesis activating neuropeptide (PBAN). We demonstrate that Plodia interpunctella is able to utilize hexadecanoic acid, and to a lesser extent tetradecanoic acid, for the biosynthesis of the main pheromone component (Z,E)-9,12-tetradecadienyl acetate. This indicated that the main pathway involves a Delta11 desaturase, chain shortening, followed by a Delta12 desaturase, but that a functional Delta9 desaturase could also be utilized. Using reverse transcription-quantitative real-time polymerase chain reaction (RT-QPCR) we distinguish two out of nine possible desaturase gene transcripts in P. interpunctella that are expressed at the highest levels. The rate-limiting step for PBAN-stimulation was studied in two moth species so as to compare the biosynthesis of a diene (P. interpunctella) and a monoene (Helicoverpa armigera) main pheromone component. In both species, incorporation of label from the (13)C sodium acetate precursor was activated by PBAN whereas no stimulatory action was observed in the incorporation of the precursors: (13)C malonyl coenzyme A; hexadecanoic 16,16,16-(2)H(3) or tetradecanoic 14,14,14-(2)H(3) acids. The acetyl coenzyme A carboxylase (ACCase) inhibitor, Tralkoxydim, inhibited the PBAN-stimulation of incorporation of stable isotope whereas the fatty-acyl reductase inhibitor, Mevastatin, failed to influence the stimulatory action of PBAN. These results provide irrefutable support to the hypothesis that PBAN affects the production of malonyl coenzyme A from acetate by the action of ACCase in the pheromone glands of these moths.