Yang_2026_Insect.Biochem.Mol.Biol_191_104555

Cholesterol homeostasis is crucial for cellular function in insects and other animals. However, the function of lipases in maintaining cholesterol homeostasis in insects was rarely investigated. In this study, we aimed to identify cholesterol-metabolizing lipases in the gut and explore their function in cholesterol regulation, using Helicoverpa armigera as a model organism. We identified 24 lipase genes containing the PF04083 domain in the H. armigera genome, four of which exhibited gut-enriched expression. Functional assays revealed that all four recombinant lipases hydrolyzed cholesterol esters, with two also capable of processing triacylglycerols (TAG). Molecular docking analyses further supported substrate specificity among these lipases. Single-gene knockout of HaOG200589 resulted in compensatory upregulation of two other lipases and cholesterol acyltransferase (ACAT). In contrast, simultaneous knockout of HaOG200551, HaOG200552, HaOG200553, and HaOG200589 led to significant downregulation of the genes coding for Niemann-Pick Type C1b (NPC1b) and ACAT. Interestingly, intestinal free cholesterol level remained relatively stable despite these gene disruption events. These findings highlight the essential role of gut-localized lipases in insect sterol metabolism.