|Ph.D Student||Zaaroor-Regev Daphna|
|Subject||A Role for the Ubiquitin System in Regulation|
of the Polycomb Group Transcriptional Repressors
Ring1B and Bmi1
|Department||Department of Medicine||Supervisor||? 18? Aaron Ciechanover|
|Full Thesis text|
The polycomb repressive complex 1 (PRC1) contains, among other components, two RING domain containing proteins Ring1B and Bmi1. Although both proteins are known repressors of the p16INK4a gene, they do not promote its ubiquitination and degradation, at least not directly. Ring1B is an E3 ubiquitin ligase that mediates its own polyubiquitination and histone H2A monoubiquitination. We found that Ring1B and Bmi1 are not targeted by their autoubiquitinating activity for degradation, but rather by an exogenous E3, independent of their RING finger domain activity. Moreover, while the RING domain of Bmi1 is required for its biological activity, Bmi1 is not a self-ubiquitinating E3. Consistent with the non-proteolytic self-ubiquitinating activity of Ring1B, it generates atypical mixed ubiquitin chains that are based on lysines 6, 27, and 48 of ubiquitin, and are required for its ability to ubiquitinate histone H2A. Importantly, we demonstrate that Bmi1 regulates Ring1B at several levels. The RING domain of both proteins mediate their association, subsequent stabilization, and probably prevent their accumulation as free, unbound PRC1 components. Independent of the stabilizing effect of Bmi1 on Ring1B, it regulates the self ubiquitinating activity of Ring1B and stimulates its histone H2A ligase activity. In the search for the E3 ligase of Ring1B we discovered that the HECT domain-containing E3 ligase, E6-AP, is able to ubiquitinate and degrade Ring1B in vitro, and in its presence Ring1B is unable to ubiquitinate histone H2A unless Bmi1 is present as well. Bmi1 also protects Ring1B from E6-AP-mediated ubiquitination and degradation. When E6-AP is absent either in cell cultures or in various tissues as brain and lung the steady state level of Ring1B increases. The elevation in Ring1B steady state level is also accompanied with an elevation in the ubiquitinated form of histone H2A and reduction of its repressed gene product HoxB9. E6-AP is mutated in the severe neurological disease, Angelman syndrome. The influence of E6-AP on Ring1B and subsequently down-stream on histone H2A and HoxB9 can imply that Ring1B has a role in the pathogenesis of Angelman syndrome and may shed new light on the events involved in brain development.