FHV genome and replicon schematics and replication strategy. (A) FHV genome is bipartite with 3.1 kb (RNA1) and 1.4 kb (RNA2) segments, and during replication a 0.4 kb subgenomic (sg) segment (RNA3) is also produced. RNA1 encodes protein A, the FHV RNA-dependent RNA polymerase (RdRp), RNA2 encodes the FHV structural protein precursor, and RNA3 encodes the RNA interference (RNAi) suppressor protein B2 (PtnB2). (B) Schematic of pS2F1, a metallothionein (MT) promoter-driven plasmid that expresses an FHV RNA1 replicon. Authentic viral 5' and 3' termini are generated by precise transcription initiation and a hepatitis δ ribozyme (δRz), respectively. The asterisk indicates the position of the frame-shifting mutation in pS2F1fs that results in the production of a truncated and non-functional RNA polymerase. Expression of the blasticidin-resistance gene (BlaR) used for selection is controlled by the copia transposon LTR constitutive promoter. (C) Strategy for FHV genome replication initiated by infection or replicon transfection. The intracellular events indicated in bold type are common for FHV RNA replication initiated via either infection or replicon induction.