Orc2 protects ORCA from ubiquitin-mediated degradation
Volume 11, Issue 19
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October 1, 2012
Pages 3578 - 3589http://dx.doi.org/10.4161/cc.21870
: Cul4A-Ddb1, ORC, ORCA/LRWD1, pre-RC, replication, ubiquitination
Authors: Zhen Shen and Supriya G. Prasanth View affiliations
Origin recognition complex (ORC) is highly dynamic, with several ORC subunits getting posttranslationally modified by phosphorylation or ubiquitination in a cell cycle-dependent manner. We have previously demonstrated that a WD repeat containing protein ORC-associated (ORCA/LRWD1) stabilizes the ORC on chromatin and facilitates pre-RC assembly. Further, ORCA levels are cell cycle-regulated, with highest levels during G1, and progressively decreasing during S phase, but the mechanism remains to be elucidated. We now demonstrate that ORCA is polyubiquitinated in vivo, with elevated ubiquitination observed at the G1/S boundary. ORCA utilizes lysine-48 (K48) ubiquitin linkage, suggesting that ORCA ubiquitination mediates its regulated degradation. Ubiquitinated ORCA is re-localized in the form of nuclear aggregates and is predominantly associated with chromatin. We demonstrate that ORCA associates with the E3 ubiquitin ligase Cul4A-Ddb1. ORCA is ubiquitinated at the WD40 repeat domain, a region that is also recognized by Orc2. Furthermore, Orc2 associates only with the non-ubiquitinated form of ORCA, and Orc2 depletion results in the proteasome-mediated destabilization of ORCA. Based on the results, we suggest that Orc2 protects ORCA from ubiquitin-mediated degradation in vivo.
Received: June 25, 2012; Accepted: August 16, 2012; Published Online: August 30, 2012
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