Lipid Droplets (LDs) were initially considered simply as a cellular energy source but are now recognised as critical organelles in signalling events, transient protein sequestration and inter-organelle interactions, however, their role in innate immune pathways, and the antiviral response remains largely unknown.
Work by our lab has demonstrated LDs are upregulated during viral infection, and that this upregulation contributes to an enhanced interferon response from the infected cell, indicating for the first time that the LD contributes to an effective immune response, however, the mechanism of this is unknown. Here, we describe for the first time that there are several critical key antiviral signalling molecules that localise to the LD during this response. We have optimised techniques to isolate pure lipid droplets from primary immortalised astrocyte cells before and following activation of viral RNA signalling pathways. Proteomic analysis has revealed there were 83 significantly upregulated proteins on LDs following stimulation with 10% of the significantly enriched proteins being associated with the interferon response. Of these, MX1, RIG-I, STAT1 and STAT2 were significantly upregulated on LD fractions at both 8 and 24hrs following RNA viral mimic stimulation. To confirm the localisation of these proteins to the LD, a technique was designed to perform fluorescent confocal microscopy on isolated lipid droplets to probe for the identified immune proteins; and this, along with western blotting, has confirmed the localisation of these proteins to LDs.
Here, we demonstrate that there are important antiviral immune signalling proteins that localise to the LDs, perhaps indicating that the LD can act as a signalling platform for signalosome formation. The mechanism by which these proteins localise to the LD and the function of this is still being explored by our laboratory.