Abstract：
Proteolysis-targeting chimeras (PROTACs) bring a protein of interest (POI) into spatial proximity of an E3 ubiquitin ligase, promoting POI ubiquitylation and proteasomal degradation. PROTACs rely on endogenous cellular machinery to mediate POI degradation, therefore the subcellular location of the POI and access to the E3 ligase being recruited potentially impacts PROTAC efficacy. To interrogate whether the subcellular context of the POI influences PROTAC-mediated degradation, we expressed either Halo or FKBP12F36V (dTAG) constructs consisting of varying localization signals and tested the efficacy of their degradation by von Hippel-Lindau (VHL)- or cereblon (CRBN)-recruiting PROTACs targeting either Halo or dTAG. POIs were localized to the nucleus, cytoplasm, outer mitochondrial membrane, endoplasmic reticulum, Golgi, peroxisome or lysosome. Differentially localized Halo or FKBP12F36V proteins displayed varying levels of degradation using the same respective PROTACs, suggesting therefore that the subcellular context of the POI can influence the efficacy of PROTAC-mediated POI degradation.
摘要：
蛋白水解靶向嵌合体（PROTACs）将感兴趣的蛋白质（POI）带入E3泛素连接酶的空间附近，促进POI泛素化和蛋白酶体降解。PROTACs依赖于内源性细胞机制来介导POI降解，因此,POI的亚细胞定位和对E3连接酶的获取可能会影响PROTAC的疗效.为了询问POI的亚细胞环境是否影响PROTAC介导的降解，我们表达了由不同定位信号组成的Halo或FKBP12F36V(dTAG)构建体,并测试了它们通过靶向Halo或dTAG的vonHippel-Lindau(VHL)-或cereblon(CRBN)-招募PROTACs降解的功效.POI定位于细胞核，细胞质，线粒体外膜，内质网，高尔基,过氧化物酶体或溶酶体。差异定位的Halo或FKBP12F36V蛋白使用相同的各自PROTACs表现出不同的降解水平，因此，表明POI的亚细胞环境可以影响PROTAC介导的POI降解的功效。
