Photoactive G-Quadruplex Ligand Identifies Multiple G-Quadruplex-Related Proteins with Extensive Sequence Tolerance in the Cellular Environment,Journal of the American Chemical Society

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Photoactive G-Quadruplex Ligand Identifies Multiple G-Quadruplex-Related Proteins with Extensive Sequence Tolerance in the Cellular Environment
Journal of the American Chemical Society ( IF 14.4 ) Pub Date : 2021-01-20 , DOI: 10.1021/jacs.0c10792
Haomiao Su , Jinglei Xu , Yuqi Chen , Qi Wang , Ziang Lu , Yage Chen _{1,

2} , Kun Chen , Shaoqing Han , Zhentian Fang , Ping Wang _{1,

2} , Bi-Feng Yuan , Xiang Zhou

Affiliation

G-Quadruplex (G4) is a noncanonical nucleic acid secondary structure with multiple biofunctions. Identifying G4-related proteins (G4RPs) is important for understanding the roles of G4 in biology. Current methods to identify G4RPs include discovery from specific biological processes or in vitro pull-down assays with specific G4 sequences. Here, we report an in vivo strategy used to identify G4RPs with extensive sequence tolerance based on G4 ligand-mediated cross-linking. Applying this method, we identified 114 and 281 G4RPs in SV589 and MM231 cells, respectively. The results successfully overlapped with all the pull-down assay literature. Through the electrophoretic mobility shift assay (EMSA), we identified some new G4-binding proteins. Moreover, enhanced cross-linking and immunoprecipitation (eCLIP) confirmed that one newly identified G4-binding protein, SERBP1, interacts with G4 in the cellular environment. The method we developed provides a new strategy for identifying proteins that interact with nucleic secondary structures in cells and benefit the study of their biological roles.

中文翻译：

光敏 G-四链体配体可识别细胞环境中具有广泛序列耐受性的多种 G-四链体相关蛋白

G-Quadruplex (G4) 是一种具有多种生物功能的非经典核酸二级结构。识别 G4 相关蛋白 (G4RP) 对于理解 G4 在生物学中的作用很重要。当前识别 G4RP 的方法包括从特定生物过程中发现或具有特定 G4 序列的体外下拉分析。在这里，我们报告了一种体内策略，用于基于 G4 配体介导的交联来识别具有广泛序列耐受性的 G4RP。应用这种方法，我们分别在 SV589 和 MM231 细胞中鉴定了 114 和 281 个 G4RP。结果成功地与所有下拉测定文献重叠。通过电泳迁移率变化测定 (EMSA)，我们鉴定了一些新的 G4 结合蛋白。而且，增强的交联和免疫沉淀 (eCLIP) 证实，一种新发现的 G4 结合蛋白 SERBP1 在细胞环境中与 G4 相互作用。我们开发的方法提供了一种新的策略，用于识别与细胞中核酸二级结构相互作用的蛋白质，并有利于对其生物学作用的研究。

更新日期：2021-01-20

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