Yeo Megan J R, Zhang Olivia, Xie Xiaowen, Nam Eunju, Payne N Connor, Gosavi Pallavi M, Kwok Hui Si, Iram Irtiza, Lee Ceejay, Li Jiaming, Chen Nicholas J, Nguyen Khanh, Jiang Hanjie, Wang Zhipeng A, Lee Kwangwoon, Mao Haibin, Harry Stefan A, Barakat Idris A, Takahashi Mariko, Waterbury Amanda L, Barone Marco, Mattevi Andrea, Carr Steven A, Udeshi Namrata D, Bar-Peled Liron, Cole Philip A, Mazitschek Ralph, Liau Brian B, Zheng Ning
Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA, USA.
Broad Institute of MIT and Harvard, Cambridge, MA, USA.
Nature. 2025 Mar;639(8053):232-240. doi: 10.1038/s41586-024-08532-4. Epub 2025 Feb 12.
UM171 is a potent agonist of ex vivo human haematopoietic stem cell self-renewal. By co-opting KBTBD4, a substrate receptor of the CUL3-RING E3 ubiquitin ligase (CRL3) complex, UM171 promotes the degradation of the LSD1-CoREST corepressor complex, thereby limiting haematopoietic stem cell attrition. However, the direct target and mechanism of action of UM171 remain unclear. Here we show that UM171 acts as a molecular glue to induce high-affinity interactions between KBTBD4 and HDAC1/2 to promote corepressor degradation. Through proteomics and chemical inhibitor studies, we identify the principal target of UM171 as HDAC1/2. Cryo-electron microscopy analysis of dimeric KBTBD4 bound to UM171 and the LSD1-HDAC1-CoREST complex identifies an asymmetric assembly in which a single UM171 molecule enables a pair of KELCH-repeat propeller domains to recruit the HDAC1 catalytic domain. One KBTBD4 propeller partially masks the rim of the HDAC1 active site, which is exploited by UM171 to extend the E3-neosubstrate interface. The other propeller cooperatively strengthens HDAC1 binding through a distinct interface. The overall CoREST-HDAC1/2-KBTBD4 interaction is further buttressed by the endogenous cofactor inositol hexakisphosphate, which acts as a second molecular glue. The functional relevance of the quaternary complex interaction surfaces is demonstrated by base editor scanning of KBTBD4 and HDAC1. By delineating the direct target of UM171 and its mechanism of action, we reveal how the cooperativity offered by a dimeric CRL3 E3 can be leveraged by a small molecule degrader.
UM171是一种体外人造血干细胞自我更新的强效激动剂。通过结合CUL3-RING E3泛素连接酶（CRL3）复合物的底物受体KBTBD4，UM171促进LSD1-CoREST共抑制复合物的降解，从而限制造血干细胞的损耗。然而，UM171的直接靶点和作用机制仍不清楚。在这里，我们表明UM171作为一种分子胶，诱导KBTBD4与HDAC1/2之间的高亲和力相互作用，以促进共抑制复合物的降解。通过蛋白质组学和化学抑制剂研究，我们确定UM171的主要靶点是HDAC1/2。对与UM171和LSD1-HDAC1-CoREST复合物结合的二聚体KBTBD4进行冷冻电子显微镜分析，确定了一种不对称组装，其中单个UM171分子使一对kelch重复螺旋桨结构域能够招募HDAC1催化结构域。一个KBTBD4螺旋桨部分掩盖了HDAC1活性位点的边缘，UM171利用这一点扩展了E3-新底物界面。另一个螺旋桨通过一个不同的界面协同加强HDAC1的结合。内源性辅因子肌醇六磷酸作为第二种分子胶，进一步加强了CoREST-HDAC1/2-KBTBD4的整体相互作用。通过对KBTBD4和HDAC1进行碱基编辑器扫描，证明了四元复合物相互作用表面的功能相关性。通过描绘UM171的直接靶点及其作用机制，我们揭示了小分子降解剂如何利用二聚体CRL3 E3提供的协同作用。
