Lysosomal acid lipase regulates cholesterol metabolism during phagosomal maturation

Abstract

Phagocytosis, a central process in innate immunity, depends on dynamic lipid remodelling, yet how cholesterol accumulates on late phagosomes remains unresolved. Here, we identify lysosomal acid lipase (LIPA) as a key cholesterol ester (CE) hydrolase driving cholesterol mobilization during phagosomal maturation. Using integrated lipidomics, chemoproteomics, and biochemical assays, we show that LIPA exhibits acidic CE hydrolase activity enriched on late phagosomes, generating free cholesterol essential for lipid raft formation. Pharmacologically inhibiting LIPA disrupts cholesterol-rich lipid raft assembly, impairs phagosomal trafficking, and alters pathogen fate — enhancing Staphylococcus aureus persistence, while restricting Mycobacterium tuberculosis survival. These findings reveal that LIPA couples cholesterol metabolism to phagocytosis, defining a mechanistic link between lipid catabolism and antimicrobial defence. By positioning CE hydrolysis as a critical determinant of phagosomal dynamics, our work uncovers a metabolic checkpoint in innate immunity and identifies LIPA as a potential therapeutic node in infection and inflammation. SIGNIFICANCE STATEMENT Phagocytosis is a central antimicrobial process, yet the metabolic signals that enable phagosomes to mature and eliminate pathogens remain poorly defined. This study uncovers lysosomal acid lipase (LIPA) as a previously unrecognized regulator of cholesterol flux during phagosomal maturation. By integrating lipidomics, chemoproteomics, and functional biochemical assays, we show that LIPA-generated cholesterol is essential for assembling lipid raft microdomains that drive the directional trafficking of late phagosomes. Disrupting this single enzymatic step impairs phagosomal progression and alters the intracellular fate of diverse pathogens, enhancing Staphylococcus aureus persistence while restricting Mycobacterium tuberculosis survival. These findings reveal that cholesterol metabolism is not merely supportive but mechanistically integral to phagosome function. By establishing LIPA as a metabolic checkpoint that couples lipid catabolism to innate immune defence, this work highlights new opportunities to modulate host–pathogen interactions through targeted manipulation of lipid pathways.