Laboratory of Genetics, The Salk Institute for Biological Studies, La Jolla, United States.
University Hospital Freiberg, University of Freiberg, Breisgau, Germany.
Elife. 2019 May 17;8:e41356. doi: 10.7554/eLife.41356.
Direct conversion of human somatic fibroblasts into induced neurons (iNs) allows for the generation of functional neurons while bypassing any stem cell intermediary stages. Although iN technology has an enormous potential for modeling age-related diseases, as well as therapeutic approaches, the technology faces limitations due to variable conversion efficiencies and a lack of thorough understanding of the signaling pathways directing iN conversion. Here, we introduce a new all-in-one inducible lentiviral system that simplifies fibroblast transgenesis for the two pioneer transcription factors, Ngn2 and Ascl1, and markedly improves iN yields. Further, our timeline RNA-Seq data across the course of conversion has identified signaling pathways that become transcriptionally enriched during iN conversion. Small molecular modulators were identified for four signaling pathways that reliably increase the yield of iNs. Taken together, these advances provide an improved toolkit for iN technology and new insight into the mechanisms influencing direct iN conversion.
直接将人类体成纤维细胞转化为诱导神经元（iNs）可以在绕过任何干细胞中间阶段的情况下产生功能性神经元。尽管 iN 技术在模拟与年龄相关的疾病以及治疗方法方面具有巨大的潜力，但由于转换效率的可变性和对指导 iN 转换的信号通路缺乏深入了解，该技术面临着限制。在这里，我们引入了一种新的一体化诱导性慢病毒系统，该系统简化了两种先驱转录因子 Ngn2 和 Ascl1 的成纤维细胞转基因，显著提高了 iN 的产量。此外，我们的 RNA-Seq 数据表明，在整个转化过程中，有信号通路在 iN 转化过程中转录丰度增加。我们鉴定了四个信号通路的小分子调节剂，这些调节剂可以可靠地增加 iNs 的产量。总之，这些进展为 iN 技术提供了一个改进的工具包，并为影响直接 iN 转换的机制提供了新的见解。
