Inhibition of potassium ion channels reduces Semliki Forest virus genome replication

Abstract

Introduction Semliki forest virus (SFV) is a model virus used to investigate the Alphavirus genus, which includes human pathogens Chikungunya virus and Ross River virus. Viruses harness cellular machinery to facilitate various steps of their replicative cycles. Ion channels are one group of cellular proteins required for the efficient replication of some viruses, including Influenza A viruses, Ebola virus and members of the Betacoronavirus genus. This study focussed on understanding SFVs requirement for functional ion channels during replication. Methods The effect of ion channel inhibitors on in vitro SFV infections was measured to investigate the contribution of ion channels in its replication cycle. Results In vitro SFV infections carried out in the presence or absence of different ion channel inhibitors showed broad-range K+ channel inhibitors reproducibly attenuated virus replication and reduced its cytotoxicity in two mammalian cell lines. These broad-range K+ channel inhibitors disrupted an early, post-entry step causing a delay or reduction in SFV protein and RNA synthesis. Screens using inhibitors of specific K+ channel families showed that two-pore domain K+ channel (2pK) inhibitors attenuated SFV replication. Confocal microscopy revealed decreased detection of dsRNA and SFV protein in the presence of inhibitor but no change in RNA and protein colocalisation, which would indicate disruption of replication complexes. Broad-range K+ and 2pK inhibitors decreased viral RNA replication and transcription from the subgenomic promoter. Conclusions K+ channel inhibitors attenuate in vitro SFV replication by inhibiting an early, post-entry step of virus replication, potentially RNA synthesis.