利用CRISPR RNA支架构建复杂的合成转录程序。
Engineering complex synthetic transcriptional programs with CRISPR RNA scaffolds.
作者信息
Zalatan Jesse G, Lee Michael E, Almeida Ricardo, Gilbert Luke A, Whitehead Evan H, La Russa Marie, Tsai Jordan C, Weissman Jonathan S, Dueber John E, Qi Lei S, Lim Wendell A
机构信息
Department of Cellular and Molecular Pharmacology, University of California San Francisco, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California San Francisco, San Francisco, CA 94158, USA.
Department of Bioengineering, University of California, Berkeley, Berkeley, CA 94720, USA; Energy Biosciences Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
出版信息
Cell. 2015 Jan 15;160(1-2):339-50. doi: 10.1016/j.cell.2014.11.052. Epub 2014 Dec 18.
Eukaryotic cells execute complex transcriptional programs in which specific loci throughout the genome are regulated in distinct ways by targeted regulatory assemblies. We have applied this principle to generate synthetic CRISPR-based transcriptional programs in yeast and human cells. By extending guide RNAs to include effector protein recruitment sites, we construct modular scaffold RNAs that encode both target locus and regulatory action. Sets of scaffold RNAs can be used to generate synthetic multigene transcriptional programs in which some genes are activated and others are repressed. We apply this approach to flexibly redirect flux through a complex branched metabolic pathway in yeast. Moreover, these programs can be executed by inducing expression of the dCas9 protein, which acts as a single master regulatory control point. CRISPR-associated RNA scaffolds provide a powerful way to construct synthetic gene expression programs for a wide range of applications, including rewiring cell fates or engineering metabolic pathways.
真核细胞执行复杂的转录程序，其中基因组中的特定基因座通过靶向调控组件以不同方式进行调控。我们已应用这一原理在酵母和人类细胞中生成基于CRISPR的合成转录程序。通过将引导RNA扩展以包括效应蛋白招募位点，我们构建了编码靶基因座和调控作用的模块化支架RNA。支架RNA集可用于生成合成多基因转录程序，其中一些基因被激活而另一些被抑制。我们应用这种方法灵活地重新引导酵母中复杂分支代谢途径的通量。此外，这些程序可通过诱导dCas9蛋白的表达来执行，dCas9蛋白作为单一的主调控控制点。与CRISPR相关的RNA支架为构建用于广泛应用的合成基因表达程序提供了一种强大的方法，包括重塑细胞命运或改造代谢途径。
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