Molecular basis of xeroderma pigmentosum group C DNA recognition by engineered meganucleases

For the gene therapy toolkit Homing endonucleases or 'meganucleases' are a type of very rare-cutting endonuclease. They recognize much larger DNA sequences than do 'classical' restriction enzymes, so they produce a low frequency of cleavage in the human genome. This makes them of interest for gene therapy, as highly specific molecular scalpels to target specific genes. In a proof-of-principle experiment, Redondo et al. have engineered homing endonucleases with the potential to aid the repair of the gene mutated in xeroderma pigmentosum, a disorder that affects the nucleotide excision repair machinery, compromising the body's ability to remove damage caused by ultraviolet light. Xeroderma pigmentosum patients have high frequencies of mutations and hence a high predisposition to develop skin cancer. The newly designed enzymes, called Amel3–4 and Ini3–4, are derivatives of the homing endonuclease I-CreI that cleave the human XPC gene both in vitro and in vivo. Structural analysis suggests that their catalytic mechanism of cleavage is similar to that of the wild type homodimeric I-CreI, and both Amel3–4 and Ini3–4 induce high levels of gene targeting in mammalian cells. This work demonstrates a gene repair technology with the potential to repair genes in xeroderma pigmentosum that produce other monogenetic diseases.