PKC phospho-activated PFK1 is required for PBAN regulated sex pheromone biosynthesis in Helicoverpa armigera

The enzyme 6-phosphofructokinase-1 (PFK1) acts as the primary rate-limiting enzyme in glycolysis, catalyzing the conversion of fructose-6-phosphate to fructose-1,6-bisphosphate. This glycolytic process provides essential substrates for the synthesis of sex pheromones. However, the specific function of PFK1 in sex pheromone biosynthesis remains unidentified. This study aimed to investigate the detailed mechanism by which PFK1 influences pheromone biosynthesis activating neuropeptide (PBAN)-regulated sex pheromone biosynthesis in Hecoverpa armigera. Findings revealed the presence of two PFK genes in pheromone glands (PGs). Further investigation demonstrated that RNAi-mediated knockdown of PFK1 significantly reduced sex pheromone production, mating success and the female ability to attract males, whereas PFK2 did not influence sex pheromone biosynthesis. Importantly, PFK1 was activated by PBAN in both isolated PGs and Sf9 cells. However, PBAN-induced activation of PFK1 could be attenuated by chelerythrine chloride (CC), a specific inhibitor of protein kinase C (PKC). Furthermore, the phosphorylation levels of PFK1 significantly increased in response to PBAN challenge, while CC treatment significantly mitigated this phosphorylation. PFK1 activity was found to depend on phosphorylation at the S135 and S676 sites in response to PBAN stimulation. Mutants at these sites abolished PFK1 phosphorylation and its activity. Overall, our findings unveil a critical mechanism by which the PBAN signaling recruits PKC to phosphorylate PFK1 at S135 and S676 sites, thereby activating PFK1. This activation ensures the normal progression of the glycolysis pathway, ultimately facilitating sex pheromone biosynthesis.