Department of Biology, Stanford University, Stanford, CA, 94305, USA.
Department of Bioengineering, Stanford University, Stanford, CA, 94305, USA.
Nat Commun. 2021 Jan 22;12(1):537. doi: 10.1038/s41467-020-20757-1.
Targeting chromatin regulators to specific genomic locations for gene control is emerging as a powerful method in basic research and synthetic biology. However, many chromatin regulators are large, making them difficult to deliver and combine in mammalian cells. Here, we develop a strategy for gene control using small nanobodies that bind and recruit endogenous chromatin regulators to a gene. We show that an antiGFP nanobody can be used to simultaneously visualize GFP-tagged chromatin regulators and control gene expression, and that nanobodies against HP1 and DNMT1 can silence a reporter gene. Moreover, combining nanobodies together or with other regulators, such as DNMT3A or KRAB, can enhance silencing speed and epigenetic memory. Finally, we use the slow silencing speed and high memory of antiDNMT1 to build a signal duration timer and recorder. These results set the basis for using nanobodies against chromatin regulators for controlling gene expression and epigenetic memory.
针对特定基因组位置的染色质调控因子的基因控制，作为一种基础研究和合成生物学中的强大方法正在出现。然而，许多染色质调控因子体积较大，难以在哺乳动物细胞中进行传递和组合。在这里，我们开发了一种使用小纳米体的基因控制策略，该纳米体可结合并募集内源性染色质调控因子至基因。我们表明，抗 GFP 纳米体可同时用于可视化 GFP 标记的染色质调控因子和控制基因表达，并且针对 HP1 和 DNMT1 的纳米体可以沉默报告基因。此外，将纳米体组合在一起或与其他调节剂（如 DNMT3A 或 KRAB）组合使用，可以增强沉默速度和表观遗传记忆。最后，我们利用抗 DNMT1 的缓慢沉默速度和高记忆性构建了信号持续时间定时器和记录器。这些结果为使用针对染色质调控因子的纳米体来控制基因表达和表观遗传记忆奠定了基础。
