Intracellular Lipid Droplet Accumulation Occurs Early Following Viral Infection and Is Required for an Efficient Interferon Response

Abstract

27Lipid droplets (LDs) are increasingly recognized as critical organelles in signalling events, 28 transient protein sequestration and inter-organelle interactions. However, the role LDs play in 29 antiviral innate immune pathways remains unknown. Here we demonstrate that induction of 30 LDs occurs as early as 2 hours post viral infection, is transient, and returns to basal levels by 31 72 hours. This phenomenon occurred following viral infections, both in vitro and in vivo. 32 Virally driven LD induction was type-I interferon (IFN) independent, however, was 33 dependent on EGFR engagement, offering an alternate mechanism of LD induction in 34 comparison to our traditional understanding of their biogenesis. Additionally, LD induction 35 corresponded with enhanced cellular type-I and -III IFN production in infected cells, with 36 enhanced LD accumulation decreasing viral replication of both HSV-1 and Zika virus 37 (ZIKV). Here, we demonstrate for the first time, that LDs play vital roles in facilitating the 38 magnitude of the early antiviral immune response specifically through the enhanced 39 modulation of IFN following viral infection, and control of viral replication. By identifying 40 LDs as a critical signalling organelle, this data represents a paradigm shift in our 41 understanding of the molecular mechanisms which coordinate an effective antiviral response. 42 48The role of LDs in an infection setting has not been well studied, however, it has been 49 demonstrated that LDs accumulate in leukocytes during inflammatory processes, and they are 50 also induced in human macrophages during bacterial infections 2 . Multiple bacterial strains, 51 including Mycobacterium spp., Chlamydia spp., Klebsiella spp. and Staphylococcus spp. are 52 known to upregulate LDs very early following bacterial infection in both primary and cell 53 culture macrophage models, and this has also been seen for a number of bacterial species in 54 rodent macrophage cell lines 5-7 . Interestingly, Trypanosoma cruzi infection of macrophages 55 also induces LDs, however, this response takes 6-12 days to occur following infection 8 . 56Bacterial-induced LD induction in immune cells has been shown to depend on toll-like 57 receptor engagement, mainly via TLR2 and TLR4, however, the role of LDs in the outcome 58 of bacterial infection remains largely unknown, and the exact mechanisms for controlling LD 59 induction remain elusive 9,10 . It has been suggested in recent work in the zebrafish model that 60 embryos with higher levels of LDs are more protected from bacterial infections 11 and work 61 in the Drosophila embryo has demonstrated that LDs can bind to histones which are released 62 upon detection of intracellular bacterial LPS and act in a bactericidal manner 12 . 63 Interestingly, LD induction has been demonstrated to be a direct result of immune activation 64 of macrophages by IFN-ߛ in a HIF-1 dependent signalling pathway 13 . M. tuberculosis 65 acquires host lipids in the absence of LDs under normal conditions, however, IFN-ߛ ...

Lipid Droplet Motility Increases Following Viral Immune Stimulation

“…To examine LD motility dynamics in the context of the activation of early innate immune pathways, we first developed a pipeline to image and track LDs in live primary immortalised astrocyte cells. Astrocytes were chosen as a model cell type, because they are known to alter LD numbers following viral stimulation and infection, and have a high number of basal LDs [ 21 ]. Additionally, astrocytes are known to be rapid producers of an effective antiviral response [ 27 ].…”

“…During viral infection, LDs have been demonstrated to be redistributed within the cell, and this has been hypothesised to be associated with viral host cell rearrangement, in order to facilitate a more effective viral life cycle [ 19 , 20 ]. However, it has recently been demonstrated that LDs are upregulated very early following both viral and bacterial infection, likely prior to pathogen replication [ 21 ] (reviewed in [ 22 ]). The dynamics such as speed and displacement of LDs remains unstudied in the context of very early viral infection; here, we demonstrate that LD dynamics significantly change following the activation of antiviral innate immune receptors.…”

“…It has now been described by us and others that LDs accumulate in pathogen-infected cells [ 21 ] (reviewed in [ 22 ]). LDs are heterogeneous, with a large spread of LD sizes ranging from < 200 to over 5000 nm (reviewed in [ 44 ]).…”

“…Pathogens such as bacteria, parasites and more recently viruses, are well known to induce LD accumulation upon host cell infection [ 21 ] (Reviewed in [ 22 ]). This accumulation of LDs has been shown to drive an enhanced immune response in respect to viral infection, and to alter LD–mitochondrial interactions which underpin metabolic changes in the cell [ 21 , 23 , 24 ]. Additionally, it has recently been demonstrated that the LD resident protein, viperin, interacts with STING, likely on the Golgi and the ER, as well as TRAF6 and IRAK1 in the TLR7/9 pathways [ 25 , 26 ].…”

Viperin interacts with PEX19 to mediate peroxisomal augmentation of the innate antiviral response

“…This raises the question as to the role of the LD in innate immune recognition of viral infection. It is emerging that the LD can act as a hub for signaling and we have recently shown that LDs influence the efficiency of the early innate response after viral infection ( Monson et al, 2018 , 2020 Preprint ). One could envisage that the LD may not be essential for MAVS-dependent signaling from the mitochondria but may be important for MAVS-dependent signaling from peroxisomes.…”