Cell and Chemical Biology, Leiden University Medical Center, Leiden, Netherlands.
Max Plank Institute for Immunobiology and Epigenetics, Freiburg, Germany.
Sci Adv. 2023 Aug 2;9(31):eadh2073. doi: 10.1126/sciadv.adh2073.
Ubiquitin and ubiquitin-like conjugation cascades consist of dedicated E1, E2, and E3 enzymes with E3s providing substrate specificity. Mass spectrometry-based approaches have enabled the identification of more than 6500 SUMO2/3 target proteins. The limited number of SUMO E3s provides the unique opportunity to systematically study E3 substrate wiring. We developed SUMO-activated target traps (SATTs) and systematically identified substrates for eight different SUMO E3s, PIAS1, PIAS2, PIAS3, PIAS4, NSMCE2, ZNF451, LAZSUL (ZNF451-3), and ZMIZ2. SATTs enabled us to identify 427 SUMO1 and 961 SUMO2/3 targets in an E3-specific manner. We found pronounced E3 substrate preference. Quantitative proteomics enabled us to measure substrate specificity of E3s, quantified using the SATT index. Furthermore, we developed the Polar SATTs web-based tool to browse the dataset in an interactive manner. Overall, we uncover E3-to-target wiring of 1388 SUMO substrates, highlighting unique and overlapping sets of substrates for eight different SUMO E3 ligases.
泛素和泛素样蛋白连接酶级联反应由专用的 E1、E2 和 E3 酶组成，其中 E3 酶提供底物特异性。基于质谱的方法已经能够鉴定出超过 6500 种 SUMO2/3 靶蛋白。有限数量的 SUMO E3 提供了系统研究 E3 底物连接的独特机会。我们开发了 SUMO 激活靶标陷阱 (SATT)，并系统地鉴定了 8 种不同 SUMO E3（PIAS1、PIAS2、PIAS3、PIAS4、NSMCE2、ZNF451、LAZSUL（ZNF451-3）和 ZMIZ2）的底物。SATT 使我们能够以 E3 特异性的方式鉴定 427 个 SUMO1 和 961 个 SUMO2/3 靶标。我们发现了明显的 E3 底物偏好。定量蛋白质组学使我们能够使用 SATT 指数测量 E3 的底物特异性。此外，我们开发了基于网络的 Polar SATTs 工具，以便以交互方式浏览数据集。总的来说，我们揭示了 1388 个 SUMO 底物的 E3 到靶标的连接，突出了 8 种不同 SUMO E3 连接酶独特和重叠的底物集合。
