Department II of Internal Medicine and Center for Molecular Medicine Cologne, University of Cologne, Faculty of Medicine and University Hospital of Cologne, Cologne, Germany.
Proteomics Core Facility, Max Planck Institute for Biology of Ageing, Cologne, Germany.
J Am Soc Nephrol. 2019 Apr;30(4):564-576. doi: 10.1681/ASN.2018090914. Epub 2019 Mar 13.
RNA-binding proteins (RBPs) are fundamental regulators of cellular biology that affect all steps in the generation and processing of RNA molecules. Recent evidence suggests that regulation of RBPs that modulate both RNA stability and translation may have a profound effect on the proteome. However, regulation of RBPs in clinically relevant experimental conditions has not been studied systematically.
We used RNA interactome capture, a method for the global identification of RBPs to characterize the global RNA-binding proteome (RBPome) associated with polyA-tailed RNA species in murine ciliated epithelial cells of the inner medullary collecting duct. To study regulation of RBPs in a clinically relevant condition, we analyzed hypoxia-associated changes of the RBPome.
We identified >1000 RBPs that had been previously found using other systems. In addition, we found a number of novel RBPs not identified by previous screens using mouse or human cells, suggesting that these proteins may be specific RBPs in differentiated kidney epithelial cells. We also found quantitative differences in RBP-binding to mRNA that were associated with hypoxia versus normoxia.
These findings demonstrate the regulation of RBPs through environmental stimuli and provide insight into the biology of hypoxia-response signaling in epithelial cells in the kidney. A repository of the RBPome and proteome in kidney tubular epithelial cells, derived from our findings, is freely accessible online, and may contribute to a better understanding of the role of RNA-protein interactions in kidney tubular epithelial cells, including the response of these cells to hypoxia.
RNA 结合蛋白（RBPs）是细胞生物学的基本调控因子，影响 RNA 分子生成和加工的所有步骤。最近的证据表明，调节既能影响 RNA 稳定性又能影响翻译的 RBP 可能对蛋白质组产生深远影响。然而，在临床相关的实验条件下，对 RBP 的调节尚未得到系统研究。
我们使用 RNA 相互作用组捕获，这是一种用于全局鉴定 RBP 的方法，以鉴定在鼠内髓集合管纤毛上皮细胞中与多聚 A 尾 RNA 种类相关的全局 RNA 结合蛋白组（RBPome）。为了研究在临床相关条件下 RBP 的调节，我们分析了缺氧相关的 RBPome 变化。
我们鉴定了 >1000 个以前使用其他系统发现的 RBP。此外，我们还发现了一些以前使用鼠或人细胞进行的筛选未鉴定的新 RBP，这表明这些蛋白质可能是分化的肾上皮细胞中特有的 RBP。我们还发现 RBP 与 mRNA 的结合在缺氧与正常氧条件下存在定量差异。
这些发现表明 RBP 受到环境刺激的调节，并深入了解了肾上皮细胞中缺氧反应信号转导的生物学。从我们的发现中得出的肾管状上皮细胞的 RBPome 和蛋白质组的知识库可在线免费获取，这可能有助于更好地理解 RNA-蛋白质相互作用在肾管状上皮细胞中的作用，包括这些细胞对缺氧的反应。
