Spielmann Malte, Kakar Naseebullah, Tayebi Naeimeh, Leettola Catherine, Nürnberg Gudrun, Sowada Nadine, Lupiáñez Darío G, Harabula Izabela, Flöttmann Ricarda, Horn Denise, Chan Wing Lee, Wittler Lars, Yilmaz Rüstem, Altmüller Janine, Thiele Holger, van Bokhoven Hans, Schwartz Charles E, Nürnberg Peter, Bowie James U, Ahmad Jamil, Kubisch Christian, Mundlos Stefan, Borck Guntram
Max Planck Institute for Molecular Genetics, 14195 Berlin, Germany; Institute for Medical Genetics and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany; Berlin-Brandenburg School for Regenerative Therapies (BSRT), 13353 Berlin, Germany;
Institute of Human Genetics, University of Ulm, 89081 Ulm, Germany; International Graduate School in Molecular Medicine Ulm, University of Ulm, 89081 Ulm, Germany; Department of Biotechnology and Informatics, BUITEMS, Quetta, 57789 Pakistan;
Genome Res. 2016 Feb;26(2):183-91. doi: 10.1101/gr.199430.115. Epub 2016 Jan 11.
The CRISPR/Cas technology enables targeted genome editing and the rapid generation of transgenic animal models for the study of human genetic disorders. Here we describe an autosomal recessive human disease in two unrelated families characterized by a split-foot defect, nail abnormalities of the hands, and hearing loss, due to mutations disrupting the SAM domain of the protein kinase ZAK. ZAK is a member of the MAPKKK family with no known role in limb development. We show that Zak is expressed in the developing limbs and that a CRISPR/Cas-mediated knockout of the two Zak isoforms is embryonically lethal in mice. In contrast, a deletion of the SAM domain induces a complex hindlimb defect associated with down-regulation of Trp63, a known split-hand/split-foot malformation disease gene. Our results identify ZAK as a key player in mammalian limb patterning and demonstrate the rapid utility of CRISPR/Cas genome editing to assign causality to human mutations in the mouse in <10 wk.
CRISPR/Cas技术能够实现靶向基因组编辑，并能快速生成用于研究人类遗传疾病的转基因动物模型。在此，我们描述了两个不相关家族中的一种常染色体隐性人类疾病，其特征为足裂缺陷、手部指甲异常和听力丧失，这是由于破坏蛋白激酶ZAK的SAM结构域的突变所致。ZAK是MAPKKK家族的成员，在肢体发育中尚无已知作用。我们发现Zak在发育中的肢体中表达，并且CRISPR/Cas介导的两种Zak异构体的敲除在小鼠胚胎期是致死性的。相比之下，SAM结构域的缺失会诱导与已知的手足裂畸形疾病基因Trp63下调相关的复杂后肢缺陷。我们的结果确定ZAK是哺乳动物肢体模式形成中的关键因子，并证明了CRISPR/Cas基因组编辑在不到10周的时间内就能快速用于在小鼠中确定人类突变的因果关系。
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