Abstract
Proper expression of the replication licensing factor Cdt1 is primarily regulated post-translationally by ubiquitylation and proteasome degradation. In a screen to identify novel non-histone targets of histone deacetylases (HDACs), we found Cdt1 as a binding partner for HDAC11. Cdt1 associates specifically and directly with HDAC11. We show that Cdt1 undergoes acetylation and is reversibly deacetylated by HDAC11. In vitro, Cdt1 can be acetylated at its N terminus by the lysine acetyltransferases KAT2B and KAT3B. Acetylation protects Cdt1 from ubiquitylation and subsequent proteasomal degradation. These results extend the list of non-histone acetylated proteins to include a critical DNA replication factor and provide an additional level of complexity to the regulation of Cdt1.
Highlights
Cdt1 levels fluctuate throughout the cell cycle
To identify potential non-histone targets of HDAC11, we made use of the fact that histone deacetylases (HDACs) often physically interact with their substrates
Specific HDAC11-interacting proteins were excised from the stained polyacrylamide gel and analyzed by mass spectrometry
Summary
Cdt1 levels fluctuate throughout the cell cycle. It is destabilized at G1/S transition, and levels begin to climb again upon S phase completion. Immunoprecipitation with anti-Myc beads followed by blotting with an anti-acetyllysine antibody showed that Myc-Cdt1 was acetylated (Fig. 2A, lane 2). Treatment of the cells with the HDAC inhibitor TSA enhances the acetylation level of Cdt1 without affecting the expression of endogenous Cdt1 (lane 2).
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