Fast type I interferon response protects astrocytes from flavivirus infection and virus-induced cytopathic effects

Lindqvist, Richard; Mundt, Filip; Gilthorpe, Jonathan D.; Wölfel, Silke; Gekara, Nelson O.; Kröger, Andrea; Överby, Anna K.

The susceptibility of IFNAR −/− astrocytes to flavivirus infection might be due to lowered basal expression of antiviral ISGs, such as viperin. Together with TRIM79α, viperin has recently been identified as an inhibitor of TBEV in vitro [50, 61] . Viperin has also been shown to have antiviral activity against WNV [62] . Viperin and TRIM79α mRNAs were quantified over time by qPCR in WT and IFNAR −/− astrocytes and WT MEFs (Fig. 5a, b) . ISGs were rapidly induced in WT astrocytes at early time points whereas induction was delayed in IFNAR −/− astrocytes and WT MEFs. The basal levels of viperin and TRIM79α were higher in WT astrocytes compared to WT MEFs and IFNAR −/− astrocytes (Fig. 5b) and might contribute to limit initial viral infection. Viperin protein was also strongly induced in WT astrocytes as early as 6 hpi whereas corresponding protein levels were only reached after 24 hpi in IFNAR −/− astrocytes (Fig. 5c) . Together, these data show that WT astrocytes express higher basal levels of a subset of antiviral genes and rapidly respond upon viral infection to express antiviral protein that can limit viral infection. MEFs and IFNAR −/− astrocytes show lower basal expression of some antiviral ISGs and a delayed IFN response and ISG were pretreated with 5000 U IFNαB/D for 16 h before infection with TBEV MOI 0.1. Viral titers were determined by focus forming assay at indicated time points (n = 6). Primary astrocytes and MEFs were infected with TBEV using MOI 0.1. Expression levels of IFN-β (c) and IFNα 2 (d) were determined by qPCR (n = 9). Supernatants were collected at the indicated time points, and antiviral activity was determined by VSV-GFP bioassay on MEFs (e, n = 6) or TBEV-based bioassay on WT and IFNAR −/− astrocytes (f, n = 12). g WT and IFNAR −/− astrocytes were treated with either 5000 U IFNαB/D or virus-inactivated supernatant from WT astrocytes 24 hpi (n= 6). TBEV replication was quantified using qPCR and normalized to input viral RNA. Mean values and standard deviations from three independent experiments are shown. *p < 0.05; **p < 0.01; ***p < 0.001 induction which is likely to render them more susceptible to viral infection.