ATRA-induced cell cycle arrest is associated with up-regulation of CKIs p21 and p27.

(A) RPA analysis of different CKIs during ATRA-induced differentiation was done using the hCC-2 multitemplate probe and total mRNA from ATRA-treated cells. (B) ATRA-induced regulation of p27 protein levels was determined by Western blot analysis using whole cell lysates of ATRA-treated cells harvested at the indicated times. (C) To determine the turnover of p27 protein, cells were grown with ATRA or left untreated, cycloheximide (CHX) was added after 72 hours of induction, and cells were harvested at 0, 1, 2, 4, 6, and 8 hours after CHX treatment. Whole cell lysates were prepared and analyzed by Western blot using antibodies directed against p27 or actin.