Xu Da, Besselink Swen, Ramadoss Gokul N, Dierks Philip H, Lubin Justin P, Pattali Rithu K, Brim Jinna I, Christenson Anna E, Colias Peter J, Ornelas Izaiah J, Nguyen Carolyn D, Chasins Sarah E, Conklin Bruce R, Nuñez James K
bioRxiv. 2024 Nov 28:2024.11.26.625496. doi: 10.1101/2024.11.26.625496.
Programmable epigenome editors modify gene expression in mammalian cells by altering the local chromatin environment at target loci without inducing DNA breaks. However, the large size of CRISPR-based epigenome editors poses a challenge to their broad use in biomedical research and as future therapies. Here, we present Robust ENveloped Delivery of Epigenome-editor Ribonucleoproteins (RENDER) for transiently delivering programmable epigenetic repressors (CRISPRi, DNMT3A-3L-dCas9, CRISPRoff) and activator (TET1-dCas9) as ribonucleoprotein complexes into human cells to modulate gene expression. After rational engineering, we show that RENDER induces durable epigenetic silencing of endogenous genes across various human cell types, including primary T cells. Additionally, we apply RENDER to epigenetically repress endogenous genes in human stem cell-derived neurons, including the reduction of the neurodegenerative disease associated V337M-mutated Tau protein. Together, our RENDER platform advances the delivery of CRISPR-based epigenome editors into human cells, broadening the use of epigenome editing in fundamental research and therapeutic applications.
可编程表观基因组编辑器通过改变目标位点的局部染色质环境来修饰哺乳动物细胞中的基因表达,而不会诱导DNA断裂。然而,基于CRISPR的表观基因组编辑器体积较大,这对它们在生物医学研究和未来治疗中的广泛应用构成了挑战。在这里,我们提出了用于将可编程表观遗传阻遏物(CRISPRi、DNMT3A-3L-dCas9、CRISPRoff)和激活剂(TET1-dCas9)作为核糖核蛋白复合物瞬时递送到人类细胞中以调节基因表达的稳健包膜递送表观基因组编辑器核糖核蛋白(RENDER)。经过合理设计,我们表明RENDER能在包括原代T细胞在内的各种人类细胞类型中诱导内源性基因的持久表观遗传沉默。此外,我们将RENDER应用于在人类干细胞衍生的神经元中表观遗传抑制内源性基因,包括减少与神经退行性疾病相关的V337M突变Tau蛋白。总之,我们的RENDER平台推动了基于CRISPR的表观基因组编辑器向人类细胞中的递送,拓宽了表观基因组编辑在基础研究和治疗应用中的使用范围。