In all eukaryotic cells, lipid droplets (LDs) store and supply essential lipids to produce signaling molecules, membrane building blocks, and metabolic energy. The LD monolayer also accommodates proteins not obviously related to lipids, such as transcription factors, chromatin components, and toxic proteins.

Common parasites (such as trypanosomes and Plasmodium falciparum), bacteria (such as mycobacteria and Chlamydia), and viruses (such as hepatitis C and dengue) induce and target LDs during their life cycles. The current view is that LDs support infection, providing microorganisms with substrates for effective growth.

Successful innate defense is critical for survival, and host species have efficiently coevolved with pathogens to develop a plethora of immune responses. Multiple cues, including cellular stress and danger-associated molecular patterns such as lipopolysaccharide (LPS), induce LD formation. Thus, LD localization and dynamics may potentially be advantageous for organizing an intracellular host defense. We have investigated the possibility that mammalian LDs have a direct and regulated role in innate immunity.

We show that mammalian LDs are endowed with a protein-mediated antimicrobial capacity, which is up-regulated during polymicrobial sepsis and by LPS. Light and electron microscopy demonstrated specific association of LDs and bacteria in human macrophages, suggesting the existence of docking mechanisms that facilitate the engagement of antibacterial LD proteins with bacteria.

A comparative mass spectrometry profiling of proteins differentially associated with LDs in response to LPS (LPS-LDs) revealed the profound remodeling of the organelle proteome. A stringent evaluation identified 689 proteins differentially regulated on LPS-LDs (317 enriched and 372 reduced). Ingenuity Pathway Analysis revealed an enrichment of innate immune system–related components and reduction of metabolism-related LD-resident proteins. Additional analyses suggested that LDs serve as innate immune hubs, integrating major intra- and extracellular immune responses.

Among the five members of the perilipin family of LD surface proteins (PLINs), PLIN5 was the only one down-regulated on LPS-LDs. PLIN5 reduction promoted physical and functional disconnection of LPS-LDs and mitochondria, with a concomitant reduction of oxidative metabolism and ketogenesis. Forced PLIN5 reexpression increased the number of LD-mitochondria contacts, reducing LD-bacteria interactions and compromising the antimicrobial capacity of cells.

By contrast, PLIN2 was the most up-regulated PLIN on LPS-LDs. Gene interaction analysis revealed that multiple immune proteins nucleated around PLIN2 in response to LPS. LPS-LDs accrued several interferon-inducible proteins such as viperin, IGTP, IIGP1, TGTP1, and IFI47. Furthermore, LPS-LDs also accumulated cathelicidin (CAMP), a broad-spectrum antimicrobial peptide with chemotactic properties. Cells overexpressing a LD-associated CAMP were more resistant to different bacterial species, including Escherichia coli, methicillin-resistant Staphylococcus aureus, and Listeria monocytogenes.

These results demonstrate that LDs form a first-line intracellular defense. They act as a molecular switch in innate immunity, responding to danger signals by both reprogramming cell metabolism and eliciting protein-mediated antimicrobial mechanisms. Mechanisms of LD trafficking and docking with phagocytic and parasitophorous membranes, observed here and described for several pathogens, may facilitate the delivery of immune proteins …