利用 CRISPR-Cas9 SunTag 系统对拟南芥基因座进行位点特异性操作。
Site-specific manipulation of Arabidopsis loci using CRISPR-Cas9 SunTag systems.
机构信息
Department of Molecular, Cell, and Developmental Biology, University of California, Los Angeles, CA, 90095, USA.
Department of Human Genetics, David Geffen School of Medicine, University of California, Los Angeles, CA, 90095, USA.
出版信息
Nat Commun. 2019 Feb 13;10(1):729. doi: 10.1038/s41467-019-08736-7.
Understanding genomic functions requires site-specific manipulation of loci via efficient protein effector targeting systems. However, few approaches for targeted manipulation of the epigenome are available in plants. Here, we adapt the dCas9-SunTag system to engineer targeted gene activation and DNA methylation in Arabidopsis. We demonstrate that a dCas9-SunTag system utilizing the transcriptional activator VP64 drives robust and specific activation of several loci, including protein coding genes and transposable elements, in diverse chromatin contexts. In addition, we present a CRISPR-based methylation targeting system for plants, utilizing a SunTag system with the catalytic domain of the Nicotiana tabacum DRM methyltransferase, which efficiently targets DNA methylation to specific loci, including the FWA promoter, triggering a developmental phenotype, and the SUPERMAN promoter. These SunTag systems represent valuable tools for the site-specific manipulation of plant epigenomes.
理解基因组功能需要通过高效的蛋白质效应子靶向系统对基因座进行特异性操作。然而，植物中可用的靶向表观基因组操作的方法很少。在这里，我们通过改造 dCas9-SunTag 系统来构建拟南芥的靶向基因激活和 DNA 甲基化工程。我们证明，利用转录激活因子 VP64 的 dCas9-SunTag 系统可以在不同的染色质环境中驱动几个基因座的强大和特异性激活，包括蛋白质编码基因和转座元件。此外，我们还提出了一种基于 CRISPR 的植物甲基化靶向系统，利用带有烟草 DRM 甲基转移酶催化结构域的 SunTag 系统，将 DNA 甲基化有效地靶向特定基因座，包括 FWA 启动子，引发发育表型，以及 SUPERMAN 启动子。这些 SunTag 系统为植物表观基因组的特异性操作提供了有价值的工具。
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