Master Regulators and Cofactors of Human Neuronal Cell Fate Specification Identified by CRISPR Gene Activation Screens.
Master Regulators and Cofactors of Human Neuronal Cell Fate Specification Identified by CRISPR Gene Activation Screens.
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DOI：
10.1016/j.celrep.2020.108460
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发表时间：
2020-12-01
期刊：
影响因子：
8.8
通讯作者：
Gersbach CA
中科院分区：
文献类型：
--
作者：
Black JB;McCutcheon SR;Dube S;Barrera A;Klann TS;Rice GA;Adkar SS;Soderling SH;Reddy TE;Gersbach CA
重新编程细胞类型规范的技术彻底改变了再生医学和疾病建模领域。目前，细胞重编程应用的命运决定因子的选择通常是一个费力且低通量的过程。因此，我们使用高通量混合 CRISPR 激活 (CRISPRa) 筛选来系统地绘制人类神经元细胞命运调节因子的图谱。我们利用基于失活 Cas9 (dCas9) 的基因激活来靶向人类基因组中的 1,496 个假定转录因子 (TF)。使用神经元承诺报告器，我们分析了这些因子在人类多能干细胞（PSC）中的神经原性活性，从而产生了一组精心设计的亲神经元因子。 TF 对的激活揭示了神经元辅助因子，包括 E2F7、RUNX3 和 LHX8，这些因子可提高转换效率、亚型特异性和神经元细胞类型的成熟。最后，利用正交 CRISPR 系统的多重基因调控，我们证明了通过同时激活和抑制靶基因来改善神经元分化，强调了基于 CRISPR 的基因调控对复杂细胞表型进行编程的能力。布莱克等人。进行混合 CRISPR 激活筛选，以确定调节人类多能干细胞神经元命运规范的因素。已确定的因素可提高转换效率并调节神经元亚型分布和成熟。总的来说，这种方法为复杂细胞表型的编程提供了一个广泛的框架。
Technologies to reprogram cell-type specification have revolutionized the fields of regenerative medicine and disease modeling. Currently, the selection of fate-determining factors for cell reprogramming applications is typically a laborious and low-throughput process. Therefore, we use high-throughput pooled CRISPR activation (CRISPRa) screens to systematically map human neuronal cell fate regulators. We utilize deactivated Cas9 (dCas9)-based gene activation to target 1,496 putative transcription factors (TFs) in the human genome. Using a reporter of neuronal commitment, we profile the neurogenic activity of these factors in human pluripotent stem cells (PSCs), leading to a curated set of pro-neuronal factors. Activation of pairs of TFs reveals neuronal cofactors, including E2F7, RUNX3, and LHX8, that improve conversion efficiency, subtype specificity, and maturation of neuronal cell types. Finally, using multiplexed gene regulation with orthogonal CRISPR systems, we demonstrate improved neuronal differentiation with concurrent activation and repression of target genes, underscoring the power of CRISPR-based gene regulation for programming complex cellular phenotypes. Black et al. perform pooled CRISPR activation screens to identify factors that regulate neuronal fate specification of human pluripotent stem cells. The identified factors improve conversion efficiencies and modulate neuronal subtype profiles and maturation. Overall, this approach provides a broad framework for programming complex cellular phenotypes.
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DOI：
10.1073/pnas.1507125112
发表时间：
2015-06-09
影响因子：
11.1
作者：
Darmanis S;Sloan SA;Zhang Y;Enge M;Caneda C;Shuer LM;Hayden Gephart MG;Barres BA;Quake SR
通讯作者：
Quake SR
影响因子：
12.3
作者：
Daley, Timothy P.;Lin, Zhixiang;Qi, Lei S.
通讯作者：
Qi, Lei S.
影响因子：
23.9
作者：
Black, Joshua B.;Adler, Andrew F.;Wang, Hong-Gang;D'Ippolito, Anthony M.;Hutchinson, Hunter A.;Reddy, Timothy E.;Pitt, Geoffrey S.;Leong, Kam W.;Gersbach, Charles A.
通讯作者：
Gersbach, Charles A.
影响因子：
5.9
作者：
Chanda, Soham;Ang, Cheen Euong;Davila, Jonathan;Pak, ChangHui;Mall, Moritz;Lee, Qian Yi;Ahlenius, Henrik;Jung, Seung Woo;Suedhof, Thomas C.;Wernig, Marius
通讯作者：
Wernig, Marius
DOI：
10.1073/pnas.1607079113
发表时间：
2016-07-26
影响因子：
11.1
作者：
Ahlenius, Henrik;Chanda, Soham;Wernig, Marius
通讯作者：
Wernig, Marius