Zinc fingers have had the golden touch in genome editing for last decade. They are the oldest, and best studied mechanism of genome modificaion. Zinc fingers (ZF) are proteins composed of conjugated Cys2His2 motifs that each recognize a specific nucleotide triplet based on the residues in their α helix.
A few years ago, this was thought to form a simple code that could be used to recognize very specific DNA sequences by engineering specific ZF motifs in tandem. By fusion of the ZF to a nuclease (i.e. a ZFN) the nuclease could cut the genome only once the ZF motifs bound the target sequence. But ZFN recognition is not so simple. The α helix residues have cross- recognition to adjacent elements, thus each motif must be chosen in the context of those around it (Liu and Stormo, 2008). These issues have now largely been addressed by better understanding the protein-DNA interactions (Cheng et al., 2012).
The FokI endonuclease that is used requires dimerization to function, meaning two custom ZNFs must bind next to each other for cutting to occur. This greatly increases the specificity. The biggest downside of ZFNs is the expense, each one is custom, and protein engineering is not a simple task each time you need to do it. ZFNs still show a lot of utility though, having recently been applied to the treatment of HIV (Tebas et al., 2014).
ZFN Additional Reading
This review gives a great overview of the history of genomic editing and ZFNs. Challenges overcome, types of genome editing now possible, and therapeutic applications are also discussed.
This review discusses ZFNs with a focus on their use in gene therapy. The basic functional mechanisms of ZFNs, design, and clinical challenges are covered.
- Cheng, L.T., Sun, L.T., and Tada, T. (2012). Genome editing in induced pluripotent stem cells. Genes Cells 17, 431-438.
- Liu, J., and Stormo, G.D. (2008). Context-dependent DNA recognition code for C2H2 zinc-finger transcription factors. Bioinformatics 24, 1850-1857.
- Tebas, P., Stein, D., Tang, W.W., Frank, I., Wang, S.Q., Lee, G., Spratt, S.K., Surosky, R.T., Giedlin, M.A., Nichol, G., et al. (2014). Gene editing of CCR5 in autologous CD4 T cells of persons infected with HIV. N. Engl. J. Med. 370, 901-910.