Gene therapy approaches to the treatment of HIV infection have targeted both viral gene expression and the cellular factors that are essential for virus replication. However, significant concerns have been raised regarding the potential toxic effects of such therapies, the emergence of resistant viral variants and unforeseen biological consequences such as enhanced susceptibility to unrelated pathogens. Novel restriction factors formed by the fusion of the tripartite motif protein (TRIM5) and cyclophilin A (CypA), or "TRIMCyps", offer an effective antiviral defence strategy with a very low potential for toxicity. In order to investigate the potential therapeutic utility of TRIMCyps in gene therapy for AIDS, a synthetic fusion protein between feline TRIM5 and feline CypA was generated and transduced into cells susceptible to infection with feline immunodeficiency virus (FIV). The synthetic feline TRIMCyp was highly efficient at preventing infection with both HIV and FIV and the cells resisted productive infection with FIV from either the domestic cat or the puma. Feline TRIMCyp and FIV infection of the cat offers a unique opportunity to evaluate TRIMCyp-based approaches to genetic therapy for HIV infection and the treatment of AIDS.