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. Author manuscript; available in PMC: 2012 Oct 7.
Published in final edited form as: Mol Cell. 2011 Oct 7;44(1):62–71. doi: 10.1016/j.molcel.2011.06.021

Figure 2. Cdt1 is stable under non-genotoxic stress conditions.

Figure 2

(A) Western blot analysis of Cdt1 expression level and chromatin binding in HeLa cells treated with sorbitol and actinomycin D. Actin and ORC2 were used as loading controls for total extract and chromatin fractions respectively.

(B) HeLa cells transfected with Flag-Cdt1 were or were not treated with sorbitol (300mM, 1 hour), and following immunoprecipitation of Flag-Cdt1, HBO1 and Geminin binding were analyzed by Western blotting.

(C) Effect of proteasome inhibition (MG132) on HBO1 association at the indicated replication origins.

(D) Western blot analysis of HA-Cdt1 immunoprecipitates in HeLa cells that were or were not treated with sorbitol. Myc-HBO1 binding, HA-Cdt1 acetylation (acetyl-K) and HA-Cdt1 phosphorylation (phospho-S/T) were investigated by Western blotting.

(E) Western blot analysis of endogenous Cdt1 immunoprecipitates for Cdt1 phosphorylation (phospho-S/T) in HeLa cells that were or were not treated with sorbitol