The molecular functions of the majority of RNA-binding proteins (RBPs) remain unclear, highlighting a major bottleneck to a full understanding of gene expression regulation. Here, we develop a plasmid resource of 690 human RBPs that we subject to luciferase-based 3ʹ-untranslated-region tethered function assays to pinpoint RBPs that regulate RNA stability or translation. Enhanced UV-cross-linking and immunoprecipitation of these RBPs identifies thousands of endogenous mRNA targets that respond to changes in RBP level, recapitulating effects observed in tethered function assays. Among these RBPs, the ubiquitin-associated protein 2-like (UBAP2L) protein interacts with RNA via its RGG domain and cross-links to mRNA and rRNA. Fusion of UBAP2L to RNA-targeting CRISPR–Cas9 demonstrates programmable translational enhancement. Polysome profiling indicates that UBAP2L promotes translation of target mRNAs, particularly global regulators of translation. Our tethering survey allows rapid assignment of the molecular activity of proteins, such as UBAP2L, to specific steps of mRNA metabolism.

大多数 RNA 结合蛋白 (RBP) 的分子功能仍不清楚，这突出了全面了解基因表达调控的主要瓶颈。在这里，我们开发了 690 个人类 RBP 的质粒资源，我们对其进行基于荧光素酶的 3'-非翻译区系链功能测定，以查明调节 RNA 稳定性或翻译的 RBP。这些 RBP 的增强 UV 交联和免疫沉淀可识别数千个响应 RBP 水平变化的内源性 mRNA 靶标，概括了在束缚功能测定中观察到的效果。在这些 RBP 中，泛素相关蛋白 2 样 (UBAP2L) 蛋白通过其 RGG 结构域与 RNA 相互作用并与 mRNA 和 rRNA 交联。UBAP2L 与靶向 RNA 的 CRISPR–Cas9 的融合展示了可编程的翻译增强。多核糖体分析表明 UBAP2L 促进目标 mRNA 的翻译，特别是全球翻译调节因子。我们的束缚调查允许将蛋白质（例如 UBAP2L）的分子活性快速分配给 mRNA 代谢的特定步骤。