Figure 1.

Strategies for combinatorial RNA interference (coRNAi). Shown is a schematic viral genome (black line) with individual RNAi target sequences (shaded boxes). Depicted are the six different possible scenarios for coRNAi discussed in detail in the text. In brief, in (a), a single region is targeted with multiple hairpin RNAs, expressed either from individual promoters (in separate or a single vector backbone(s)) or as concatemers from one promoter. Another possibility, not mentioned in the text, to increase hairpin RNA levels is the use of an inherently stronger promoter (thicker arrow). Similarly, constructs expressing multiple small hairpin RNAs (shRNAs) from one or separate promoters can be designed simultaneously to target various (b) viral or (c) combinations of viral and cellular genes. (d) A unique means of co-silencing numerous regions on a single target is expression of a long hairpin RNA from one promoter. Recently, coRNAi vectors were refined to co-express short hairpin RNAs together with other nucleic acid–based inhibitors (ribozymes (Rzs) and HIV trans-activation response region (TAR) decoy (e)) or with transdominant ((f), gray circles; the asterisk indicates point mutations)) or therapeutic proteins (f, black circle).