Nucleosome Acidic Patch Promotes RNF168- and RING1B/BMI1-Dependent H2AX and H2A Ubiquitination and DNA Damage Signaling

摘要

Histone ubiquitinations are critical for the activation of the DNA damage response (DDR). In particular, RNF168 and RING1B/BMI1 function in the DDR by ubiquitinating H2A/H2AX on Lys-13/15 and Lys-118/119, respectively. However, it remains to be defined how the ubiquitin pathway engages chromatin to provide regulation of ubiquitin targeting of specific histone residues. Here we identify the nucleosome acid patch as a critical chromatin mediator of H2A/H2AX ubiquitination (ub). The acidic patch is required for RNF168- and RING1B/BMI1-dependent H2A/H2AXub in vivo . The acidic patch functions within the nucleosome as nucleosomes containing a mutated acidic patch exhibit defective H2A/H2AXub by RNF168 and RING1B/BMI1 in vitro . Furthermore, direct perturbation of the nucleosome acidic patch in vivo by the expression of an engineered acidic patch interacting viral peptide, LANA, results in defective H2AXub and RNF168-dependent DNA damage responses including 53BP1 and BRCA1 recruitment to DNA damage. The acidic patch therefore is a critical nucleosome feature that may serve as a scaffold to integrate multiple ubiquitin signals on chromatin to compose selective ubiquitinations on histones for DNA damage signaling. Author Summary Post-translational modifications of histones play important roles in regulating both the structure and function of chromatin. As all DNA based processes, including transcription, DNA replication and DNA repair, occur within the context of chromatin, the actual in vivo substrate of these reactions is chromatin. Thus, understanding these processes within the context of chromatin is vital for providing mechanistic insights into chromatin-based processes, including DNA damage signaling and genome maintenance. Here we identify a structure within H2A and H2AX termed the acidic patch that promotes the activity of two independent ubiquitin E3 ligase complexes, RNF168 and RING1B/BMI1, and is required for DNA damage ubiquitin signaling. We show directly in vitro and in vivo that this nucleosome structure is critical for histone H2A and H2AX ubiquitinations and the DNA damage response in cells. In addition, we engineered a novel biological tool that blocked the nucleosome acidic patch of all histone H2A species leading to the repression of the DNA damage response in cells. Collectively, DNA damage factors elicit their response not only through histone modifications such as ubiquitin but also through interactions within nucleosome surface structures to activate DNA damage signaling.