Elucidating the RNA–Protein Interactomes of Target RNAs in Tissue

RNA–protein interactions are key to many aspects
of cellular
homeostasis and their identification is important to understanding
cellular function. Multiple strategies have been developed for the
RNA-centric characterization of RNA–protein complexes. However,
these studies have all been done in immortalized cell lines that do
not capture the complexity of heterogeneous tissue samples. Here,
we develop hybridization purification of RNA–protein complexes
followed by mass spectrometry (HyPR-MS) for use in tissue samples.
We isolated both polyadenylated RNA and the specific long noncoding
RNA MALAT1 and characterized their protein interactomes. These results
demonstrate the feasibility of HyPR-MS in tissue for the multiplexed
characterization of specific RNA–protein complexes.

RNA-蛋白质相互作用（RNA–protein interactions）是细胞稳态诸多环节的核心调控因素，对其进行鉴定对于理解细胞功能具有重要意义。目前已开发出多种以RNA为中心的RNA-蛋白质复合物表征策略，但此类研究均基于永生化细胞系开展，无法还原异质性组织样本的真实复杂性。本研究开发了适用于组织样本的杂交纯化结合质谱RNA-蛋白质复合物分析技术（hybridization purification of RNA–protein complexes followed by mass spectrometry，HyPR-MS）。我们分别分离了聚腺苷酸化RNA与特异性长链非编码RNA（long noncoding RNA）MALAT1，并对二者的蛋白质相互作用组进行了表征。研究结果证实，HyPR-MS可用于组织样本中特定RNA-蛋白质复合物的多重表征，具备实际应用可行性。