PIE-seq: identifying RNA-binding protein targets by dual RNA-deaminase editing and sequencing.
PIE-seq: identifying RNA-binding protein targets by dual RNA-deaminase editing and sequencing.
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DOI：
10.1038/s41467-023-39054-8
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发表时间：
2023-06-06
影响因子：
16.6
通讯作者：
Zhang, Xiaochang
中科院分区：
文献类型：
--
作者：
Ruan, Xiangbin;Hu, Kaining;Zhang, Xiaochang
RNA结合蛋白(RBPs)对于基因调控是必不可少的，但确定其跨细胞类型的RNA靶标仍然是一个挑战。在这里，我们提出了PIE-Seq，通过将C-to-U和A-to-I碱基编辑与限制性商业惯例结合起来，利用双重脱氨酶编辑和测序来研究蛋白质-RNA的相互作用。我们对PIE-Seq进行了基准测试，并展示了它在单细胞中的敏感性、它在发育中的大脑中的应用以及它以25个人类RBP为例的可扩展性。Bulk PIE-Seq识别限制性商业惯例的典型结合特征，如PUM2和NOVA1，并为大多数测试的限制性商业惯例提名额外的靶基因，如SRSF1和TDP-43/TARDBP。同源限制性片断经常编辑PIE-Seq中相似的序列和基因集，而不同的RBP家族显示不同的靶点。单细胞PIE-PUM2发现了与大样本类似的靶点，并将PIE-PUM2应用于发育中的小鼠新皮质，确定了神经前体和神经元特异性靶基因，如App。总之，PIE-Seq提供了一种正交的方法和资源来发现小鼠和人类细胞中的RBP靶标。追踪不同细胞类型的蛋白质-RNA相互作用是具有挑战性的。在这里，Ruan等人开发了一种称为PIE-Seq的双重脱氨酶方法，其中蛋白质靶标由C-to-U和A-to-I RNA碱基编辑标记，并将其应用于25种人类RNA结合蛋白质。
RNA-binding proteins (RBPs) are essential for gene regulation, but it remains a challenge to identify their RNA targets across cell types. Here we present PIE-Seq to investigate Protein-RNA Interaction with dual-deaminase Editing and Sequencing by conjugating C-to-U and A-to-I base editors to RBPs. We benchmark PIE-Seq and demonstrate its sensitivity in single cells, its application in the developing brain, and its scalability with 25 human RBPs. Bulk PIE-Seq identifies canonical binding features for RBPs such as PUM2 and NOVA1, and nominates additional target genes for most tested RBPs such as SRSF1 and TDP-43/TARDBP. Homologous RBPs frequently edit similar sequences and gene sets in PIE-Seq while different RBP families show distinct targets. Single-cell PIE-PUM2 uncovers comparable targets to bulk samples and applying PIE-PUM2 to the developing mouse neocortex identifies neural-progenitor- and neuron-specific target genes such as App. In summary, PIE-Seq provides an orthogonal approach and resource to uncover RBP targets in mice and human cells. Tracking protein-RNA interaction across cell types is challenging. Here, Ruan et al develop a dual-deaminase method called PIE-Seq, where protein targets are marked by both C-to-U and A-to-I RNA base editors, and apply it to 25 human RNA-binding proteins.
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影响因子：
64.5
作者：
Cooper TA;Wan L;Dreyfuss G
通讯作者：
Dreyfuss G
影响因子：
64.5
作者：
Hafner M;Landthaler M;Burger L;Khorshid M;Hausser J;Berninger P;Rothballer A;Ascano M Jr;Jungkamp AC;Munschauer M;Ulrich A;Wardle GS;Dewell S;Zavolan M;Tuschl T
通讯作者：
Tuschl T
影响因子：
3.6
作者：
Das AT;Tenenbaum L;Berkhout B
通讯作者：
Berkhout B
DOI：
10.1016/j.omtn.2021.07.008
发表时间：
2021-09-03
期刊：
Molecular therapy. Nucleic acids
影响因子：
--
作者：
Kluesner MG;Tasakis RN;Lerner T;Arnold A;Wüst S;Binder M;Webber BR;Moriarity BS;Pecori R
通讯作者：
Pecori R
影响因子：
14.9
作者：
Bohn, Jennifer A.;Van Etten, Jamie L.;Goldstrohm, Aaron C.
通讯作者：
Goldstrohm, Aaron C.