Figure 1: Cdc20 is phosphorylated in interphase.
From: Interphase APC/C–Cdc20 inhibition by cyclin A2–Cdk2 ensures efficient mitotic entry
(a) Schematic of Cdc20 with known Cdk sites indicated by red circles and residue number. The C-Box motif, WD40 domain and IR tail is indicated as well. (b) YFP-tagged Cdc20, Cdc20 T59A or Cdc20 T70A was transiently expressed in HeLa cells and isolated using an YFP affinity resin from nocodazole-arrested cells harvested by mitotic shake-off. The purified complexes were analysed by SDS–PAGE and probed with polyclonal phospho-specific antibodies against Cdc20 T59 and Cdc20 T70, as well as a pan-specific Cdc20 antibody. (c) Synchronization protocol used to perform a double thymidine block and release experiment. D1–4 indicates day 1–4. (d,e) Western blot of input and Cdc20-purified samples from HeLa cells prepared as indicated in c and probed with the indicated antibodies. Representative of at least two independent experiments. (f,g) Western blot of APC3 and BubR1 purified samples prepared as in c. Representative of at least two independent experiments.
