Figure 2

Turning a flavivirus into a viral vector for therapeutic small RNA delivery. a) The flavivirus genome consists of a 5′ untranslated region (5′UTR), followed by an open reading frame (encoding the structural and non-structural proteins as a polyprotein), and finishing with the 3′UTR. Some cis-acting RNA secondary structures that participate in virus replication and translation are depicted. b) The first step during generation of a flavivirus-based viral vector is the construction of a flavivirus replicon by reverse genetics, in which the excision/replacement of the structural genes by the reporter/selectable marker (green), and the insertion of the DNA encoding a shRNAmir or artificial miRNA precursor in the hypervariable region of the 3′UTR can be performed. c) Viral structural genes can be expressed in trans from an expression plasmid or a packaging cell line generated by selection of the transfected cells, ensuring biosafety by allowing replication but avoiding propagation of the flavivirus vector after transduction of the target cells.