Native homing endonucleases can target conserved genes in humans and in animal models

Adi Barzel, Eyal Privman, Michael Peeri, Adit Naor, Einat Shachar, David Burstein, Rona Lazary, Uri Gophna, Tal Pupko, Martin Kupiec

Research output: Contribution to journalArticlepeer-review

Abstract

In recent years, both homing endonucleases (HEases) and zinc-finger nucleases (ZFNs) have been engineered and selected for the targeting of desired human loci for gene therapy. However, enzyme engineering is lengthy and expensive and the off-target effect of the manufactured endonucleases is difficult to predict. Moreover, enzymes selected to cleave a human DNA locus may not cleave the homologous locus in the genome of animal models because of sequence divergence, thus hampering attempts to assess the in vivo efficacy and safety of any engineered enzyme priorto its application in human trials. Here, we show that naturally occurring HEases can be found, that cleave desirable human targets. Some of these enzymes are also shown to cleave the homologous sequence in the genome of animal models. In addition, the distribution of off-target effects may be more predictable for native HEases. Based on our experimental observations, we present the HomeBase algorithm, database and web server that allow a high-throughput computational search and assignment of HEases for the targeting of specific loci in the human and other genomes. We validate experimentally the predicted target specificity of candidate fungal, bacterial and archaeal HEases using cell free, yeast and archaeal assays.

Original languageEnglish
Pages (from-to)6646-6659
Number of pages14
JournalNucleic Acids Research
Volume39
Issue number15
DOIs
StatePublished - Aug 2011

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