Study reveals the molecular mechanisms of dynamic activation of DNMT1 by H3Ub2

Reviewed by Emily Henderson, B.Sc.Oct 28 2022

This study is led by Dr. Na Yang (State Key Laboratory of Medicinal Chemical Biology, College of Pharmacy and Key Laboratory of Medical Data Analysis and Statistical Research of Tianjin, Nankai University), and Dr. Bing Zhu (National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Science). Dr. Jixue Sun is the first author of this article.

Previous studies showed that ubiquitin-like nuclear cofactor UHRF1 activates DNMT1 in a histone H3 ubiquitination-dependent manner (Bostick et al., 2007; Nishiyama et al., 2013; Qin et al., 2015; Sharif et al., 2007). H3Ub2 recognizes DNMT1 by binding to the RFTS domain. The resolved structure of the H3Ub2-RFTS complex shows that histone H3 occupies the position of an auto-inhibitory loop of DNMT1, and the two ubiquitin groups intimately contact with the N-lobe of RFTS. Compared with the apo structure of RFTS, the α4-helix undergoes an obvious counter clockwise bend (Ishiyama et al., 2017; Li et al., 2018), which is thought to be the key to activate DNMT1 with further conformational changes. However, the dynamic mechanism of the activation of DNMT1 by H3Ub2 has not been completely elucidated at the molecular level because of the lack of entire structural information, or spatial and temporal resolution limitations of previous studies.

In the study, it was found that H3Ub2 promotes the bending of the α4-helix of RFTS during the multi-scale molecular dynamics simulations, and the "bending" conformation can be induced back to a "straight" conformation when H3Ub2 is removed from the simulation systems. In addition, the results of Principal Component analysis combined with Community Cluster analysis showed that the RFTS domain rotates ~20° anti-clockwise and moves ~3 Å away from the target recognition domain (TRD) upon H3Ub2 binding, resulting in the exposure of the catalytic core of DNMT1.

Furthermore, the hydrogen-bonding network at the interfaces of RFTS-TRD and RFTS-CD is significantly disrupted after H3Ub2 binding. Alanine substitutions of all the residues crucial for hydrogen bond formation weaken the interactions between the RFTS domain and catalytic core of DNMT1, resulting in large and fast conformational changes during the simulations, which contributes to activating the auto-inhibited conformation of DNMT1 by opening the substrate binding pocket in CD.

This study revealed the molecular mechanisms of the dynamic activation of DNMT1 by H3Ub2, including: ① Inducing the bending of the α4-helix in the RFTS domain of DNMT1. ② Promoting the rotation of RFTS away from the TRD domain close to the catalytic core. ③Attenuating the interactions between RFTS and TRD-CD that facilitates the exposing of DNA binding pocket in CD.