Native homing endonucleases can target conserved genes in humans and in animal models.
Native homing endonucleases can target conserved genes in humans and in animal models.
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DOI：
10.1093/nar/gkr242
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发表时间：
2011-08
影响因子：
14.9
通讯作者：
Kupiec M
中科院分区：
文献类型：
--
作者：
Barzel A;Privman E;Peeri M;Naor A;Shachar E;Burstein D;Lazary R;Gophna U;Pupko T;Kupiec M
近年来，归巢核酸内切酶（HEases）和锌指核酸酶（ZFN）都已被工程化并被选择用于靶向用于基因治疗的所需人类基因座。然而，酶工程是冗长和昂贵的，并且所制造的核酸内切酶的脱靶效应难以预测。此外，由于序列差异，选择用于切割人DNA基因座的酶可能不切割动物模型基因组中的同源基因座，因此阻碍了在其应用于人体试验之前评估任何工程化酶的体内功效和安全性的尝试。在这里，我们表明可以发现天然存在的HEase，其切割所需的人类靶标。这些酶中的一些也显示出切割动物模型基因组中的同源序列。此外，脱靶效应的分布对于天然HEase可能更可预测。根据我们的实验观察，我们提出了基于算法，数据库和Web服务器，允许高通量的计算搜索和分配HEases在人类和其他基因组中的特定位点的目标。我们使用无细胞、酵母和古细菌测定实验验证候选真菌、细菌和古细菌HEase的预测靶特异性。
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 prior to 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.
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影响因子：
5.3
作者：
Aylon, Y;Liefshitz, B;Kupiec, M
通讯作者：
Kupiec, M
影响因子：
46.9
作者：
Hockemeyer, Dirk;Soldner, Frank;Beard, Caroline;Gao, Qing;Mitalipova, Maisam;DeKelver, Russell C.;Katibah, George E.;Amora, Ranier;Boydston, Elizabeth A.;Zeitler, Bryan;Meng, Xiangdong;Miller, Jeffrey C.;Zhang, Lei;Rebar, Edward J.;Gregory, Philip D.;Urnov, Fyodor D.;Jaenisch, Rudolf
通讯作者：
Jaenisch, Rudolf
影响因子：
20.3
作者：
Cavazzana-Calvo, Marina;Carlier, Frederique;Hacein-Bey-Abina, Salima
通讯作者：
Hacein-Bey-Abina, Salima
影响因子：
14.9
作者：
Brok-Volchanskaya, Vera S.;Kadyrov, Farid A.;Granovsky, Igor E.
通讯作者：
Granovsky, Igor E.
影响因子：
5.6
作者：
Edgell, DR;Stanger, MJ;Belfort, M
通讯作者：
Belfort, M