Department of Molecular Medicine, Scripps Research, 10550 N Torrey Pines Rd, La Jolla, California 92037, United States.
Department of Genetics, Stanford University, Stanford, California 94305, United States.
J Proteome Res. 2022 Sep 2;21(9):2197-2210. doi: 10.1021/acs.jproteome.2c00281. Epub 2022 Aug 16.
Emerin and lamin B receptor (LBR) are abundant transmembrane proteins of the nuclear envelope that are concentrated at the inner nuclear membrane (INM). Although both proteins interact with chromatin and nuclear lamins, they have distinctive biochemical and functional properties. Here, we have deployed proximity labeling using the engineered biotin ligase TurboID (TbID) and quantitative proteomics to compare the neighborhoods of emerin and LBR in cultured mouse embryonic fibroblasts. Our analysis revealed 232 high confidence proximity partners that interact selectively with emerin and/or LBR, 49 of which are shared by both. These included previously characterized NE-concentrated proteins, as well as a host of additional proteins not previously linked to emerin or LBR functions. Many of these are TM proteins of the ER, including two E3 ubiquitin ligases. Supporting these results, we found that 11/12 representative proximity relationships identified by TbID also were detected at the NE with the proximity ligation assay. Overall, this work presents methodology that may be used for large-scale mapping of the landscape of the INM and reveals a group of new proteins with potential functional connections to emerin and LBR.
出核蛋白（emerin）和核膜内层（INM）受体（lamin B receptor，LBR）是核膜上丰富的跨膜蛋白，它们在内核膜处聚集。尽管这两种蛋白都与染色质和核纤层相互作用，但它们具有独特的生化和功能特性。在这里，我们使用工程化的生物素连接酶 TurboID（TbID）进行邻近标记，并结合定量蛋白质组学，比较了培养的小鼠胚胎成纤维细胞中出核蛋白和 LBR 的邻近区域。我们的分析揭示了 232 个高可信度的邻近伴侣，它们与出核蛋白和/或 LBR 选择性相互作用，其中 49 个与两者都有关。这些伴侣包括先前表征的核周集中蛋白，以及许多以前与出核蛋白或 LBR 功能无关的额外蛋白。其中许多是内质网（ER）的跨膜蛋白，包括两种 E3 泛素连接酶。支持这些结果，我们发现 TbID 鉴定的 11/12 个代表性邻近关系也在用邻近连接检测法在 NE 处被检测到。总的来说，这项工作提出了一种可用于大规模绘制 INM 景观的方法，并揭示了一组具有与出核蛋白和 LBR 潜在功能联系的新蛋白。
