ABSTRACT
Zinc finger nucleases (ZFN) are hybrid molecules composed of a designed polymeric zinc finger domain specific for a DNA target sequence and a FokI nuclease cleavage domain. FokI requires dimerization to expurgate DNA. The binding of two heterodimers of designed ZFN-FokI hybrid molecules to two adjoining target sequences in each DNA strand disjointed by a 6 base-pair cleavage site results in FokI dimerization and ensuing DNA cleavage. The ZFN architecture is said to be meeting these specifications by joining the DNA-binding domain of a versatile class of eukaryotic transcription features called zinc finger proteins (ZFPs) with the nuclease domain of the FokI restriction enzyme. Generation of a DNA double-strand break (DSB) by a ZFN results in the instigation of a cellular response well-known as the DNA damage response. ZFNs technique has been used to disrupt native loci in model organisms such as rats and Arabidopsis thaliana, to drive trait stacking in a crop species, to engineer HIV-resistant human T cells and hematopoietic stem cells (HSCs), and to drive targeted integration in human embryonic stem (ES) cells, induced pluripotent stem (iPS) cells, and mesenchymal stem cells. ZFNs can be used to edit genomes of numerous cell types and organisms. ZFNs have gene modification efficiencies of around 29%. There are about 30 types of ZNFs that are approved by the HUGO Gene Nomenclature Committee. The classification is based on the structure of the zinc finger domain. The applications of genome editing employing ZFNs are grounded on the introduction of a site-specific DNA DSB into the locus of concern. ZFN-stimulated gene targeting can preserve temporal and tissue-specific gene expression better than conventional gene-addition-type gene therapy. ZFNs have complex requirements in designing and constructing them, resulting in a high failure rate. They have a limited application in high throughput screening. ZFNs were used to efficiently and specifically excise HIV-1 proviral DNA, and this was achieved in latently infected human T cells.
