CDC6 expression is coupled to phosphorylation of CDK2 on Thr-160 and
exit from quiescence induced by serum. A, BJ-hTERT cells were
transfected with siRNAs targeting CDC6 or renilla luciferase in conditions of
serum starvation. Forty-eight hours later, cells were restimulated with 10%
FBS, and 48 h after restimulation, cells were harvested and lysed for
soluble/chromatin-bound protein fractionation and Western blot analysis.
B and C, BJ-hTERT cells were transfected with siRNAs
targeting CDC6 or EGFP in conditions of serum starvation. Seventy-two hours
later, cells were re-stimulated with FBS and harvested at the indicated time
points. Cell pellets were divided in two for soluble/chromatin-bound protein
fractionation and Western blot analysis (B) and PI/FACS analysis
(C). A and B, same exposure times are shown for
both fractions. B, CDK2 band corresponding to phosphorylated Thr-160
is indicated with an asterisk. LE indicates long exposure.
SE indicates short exposure. C, percent of cells in each
cell cycle phase is indicated.