The flavivirus genus includes important human pathogens like Tick-borne encephalitis virus (TBEV), Dengue virus (DV) and West-Nile virus (WNV), that can cause severe disease e.g. encephalitis or hemorrhagic fever. The NS5 protein is a multifunctional RNA dependent RNA polymerase indispensable for the flavivirus replication. We have previously shown that TBEVNS5 contains a unique internal PDZ binding motif (YS223) for specific targeting of the PDZ protein Scribble. This interaction has impact on both viral down regulation of host cellular defense systems and neurite outgrowth. Putative C-terminal PDZ binding motifs present in TBEVNS5 (-SII903) and WNVNS5 (-TVL905) have also previously been highlighted.
To determine whether the PDZ binding motifs of TBEVNS5 has an effect on virus replication we constructed a DNA based sub-genomic TBEV replicon expressing firefly luciferase. The motifs within NS5 were mutated individually and in concert and the replicons were assayed in cell culture. Our results show that the replication rate was impaired in all mutants, which indicates that PDZ dependent host interactions influence flavivirus replication.We also find that the C-terminal PDZ binding motif present in TBEVNS5 and WNVNS5 are targeting various human PDZ domain proteins. TBEVNS5 has high affinity to Zonulaoccludens-2 (ZO-2),GIAP C-terminus interacting protein (GIPC), Calcium/calmodulin-dependent serine protein kinase (CASK) and Interleukin 16 (IL-16).A different pattern was observed for WNVNS5 as it associated with IL-16, and several other putative interaction partners.