Large-scale map of RNA binding protein interactomes across the mRNA life-cycle (Street, Rothamel, and Brannan et al., 2024)

mRNAs interact with RNA-binding proteins (RBPs) throughout their processing and maturation. While substantial effort has assigned RBPs to RNA substrates, less exploration has leveraged protein-protein interactions (PPIs) to study proteins in mRNA life-cycle stages. We generated an RNA-aware RBP-centric PPI map across the mRNA life-cycle by immunopurification-mass spectrometry of ~100 endogenous RBPs in the presence or absence of RNase, augmented by size exclusion chromatography-mass spectrometry. We identify 8,742 known and 20,802 novel interactions between 1125 proteins and determine that 73% of IP interactions are RNA-regulated. Our interactome links proteins, many of unknown function, with distinct life-cycle stages, linking nearly half the proteins to multiple stages. We demonstrate the value of this resource by assigning novel functions to the two most connected proteins. We show that ERH functions in multiple stages, including splicing and export, and that splicing protein SNRNP200 interacts with stress granule proteins and binds cytoplasmic RNA differently during stress

信使RNA在其加工和成熟过程中与RNA结合蛋白（RBPs）相互作用。尽管大量研究将RBPs分配到RNA底物中，但利用蛋白质-蛋白质相互作用（PPIs）来研究mRNA生命周期阶段中的蛋白质的研究相对较少。我们通过免疫纯化-质谱法对约100种内源性RBPs进行免疫纯化，在存在或不存在RNase的情况下，结合尺寸排阻色谱-质谱法，生成了一种RNA感知的以RBPs为中心的PPI图谱，覆盖了mRNA的生命周期。我们鉴定出1125种蛋白质之间存在的8742个已知和20802个新相互作用，并确定73%的免疫沉淀相互作用受RNA调控。我们的互作组将许多功能未知的蛋白质与不同的生命周期阶段联系起来，将近一半的蛋白质与多个阶段相联系。我们通过为两个最连接的蛋白质赋予新功能，展示了这一资源的价值。我们发现ERH在多个阶段发挥作用，包括剪接和输出，以及剪接蛋白SNRNP200与应激颗粒蛋白相互作用，并在应激状态下以不同的方式结合细胞质RNA。