Species-specific sex pheromones released by female moths to attract conspecific male moths are synthesized de novo in the pheromone gland (PG) via the fatty acid biosynthetic pathway. This pathway is regulated by a neurohormone termed pheromone biosynthesis activating neuropeptide (PBAN), a 33-amino acid peptide that originates in the subesophageal ganglion. In the silkmoth, Bombyx mori, cytoplasmic lipid droplets, which store the sex pheromone (bombykol) precursor fatty acid, accumulate in PG cells. PBAN stimulates lipolysis of the stored lipid droplet triacylglycerols (TAGs) and releases the precursor for final modification. PBAN exerts its physiological function via the PG cell-surface PBAN receptor, a G protein-coupled receptor that belongs to the neuromedin U receptor family. The PBAN receptor-mediated signal is transmitted via a canonical store-operated channel activation pathway utilizing Gq-medi- Mating in moths is limited to a specific phase of the photoperiod and developmental stage. Accordingly, the biochemical processes that comprise sex pheromone biosynthesis in female moths must be precisely regulated. In most moth species these processes are regulated by a neurohormone termed pheromone biosynthesis activating neuropeptide (PBAN), 4 a 33-amino acid peptide that originates in the subesophageal ganglion and that is characterized by a core C-terminal FSPRLamide sequence (1, 2). After adult emergence, PBAN is released into the hemolymph during a species-specific period and acts on the pheromone gland (PG) to trigger the production and release of species-specific sex pheromones (3, 4).PG is a functionally differentiated organ in close proximity to the terminal abdominal tip that originates in the intersegmental membrane between the 8th and 9th abdominal segments (5-7). In the silkmoth, Bombyx mori, the sex pheromone, E,Z-10,12-hexadecadien-1-ol, commonly known as bombykol, is synthesized de novo within PG cells from acetyl-CoA via the conventional long chain fatty acid biosynthetic pathway (8, 9). The straight chain fatty acyl intermediate, palmitate, is converted stepwise to bombykol by the actions of a bifunctional Z11-10/12 fatty acyl desaturase, Bmpgdesat1, and a PG-specific fatty acyl reductase, pgFAR (10 -12). On the day before adult emergence, B. mori PG cells rapidly accumulate numerous lipid droplets (LDs) within the cytoplasm (13). These LDs play an essential role in bombykol biosynthesis by acting as a reservoir for the de novo synthesized bombykol precursor, ⌬10,12-hexadecadienoate, which is deposited in the LDs in the form of triacylglycerols (TAGs) with the precursor predominantly sequestered at the sn-1 and sn-3 positions of the glycerides (14). After adult emergence, the stored fatty acid is cleaved and converted to bombykol in response to PBAN (7,15).The pheromonotropic effects of PBAN are dependent on extracellular Ca 2ϩ (3,4) and are mediated by the PG cell-surface PBAN receptor, a G protein-coupled receptor that belongs to the neuromedin U receptor family (16 -18

Species-specific sex pheromones released by female moths to attract conspecific male moths are synthesized de novo in the pheromone gland (PG) via fatty acid synthesis (FAS). Biosynthesis of moth sex pheromones is usually regulated by a neurohormone termed pheromone biosynthesis activating neuropeptide (PBAN), a 33-aa peptide that originates in the subesophageal ganglion. In the silkmoth, Bombyx mori, cytoplasmic lipid droplets (LDs), which store the sex pheromone (bombykol) precursor fatty acid, accumulate in PG cells prior to eclosion. PBAN activation of the PBAN receptor stimulates lipolysis of the stored LD triacylglycerols (TAGs) resulting in release of the bombykol precursor for final modification. While we have previously characterized a number of molecules involved in bombykol biosynthesis, little is known about the mechanisms of PBAN signaling that regulate the TAG lipolysis in PG cells. In the current study, we sought to further identify genes involved in bombykol biosynthesis as well as PBAN signaling, by using a subset of 312 expressed-sequence tag (EST) clones that are in either our B. mori PG cDNA library or the public B. mori EST databases, SilkBase and CYBERGATE, and which are preferentially expressed in the PG. Using RT-PCR expression analysis and an RNAi screening approach, we have identified another eight EST clones involved in bombykol biosynthesis. Furthermore, we have determined the functional role of a clone designated BmACP that encodes B. mori acyl carrier protein (ACP). Our results indicate that BmACP plays an essential role in the biosynthesis of the bombykol precursor fatty acid via the canonical FAS pathway during pheromonogenesis.