miR-34a regulates CDK4 and CDK6 in K562 cells. (A) K562 cells, transfected with the indicated miRNA mimic, were analyzed for CDK4 and CDK6 protein and mRNA by immunoblot (left) or quantitative PCR (right), respectively, 48 hours later. Protein expression relative to α-tubulin was quantified by densitometry. mRNA normalized to GAPDH mRNA is shown relative to the ratio in K562 cells transfected with a miRNA mimic control. (B) Simultaneous knockdown (top left) of CDK4 and CDK6, but not either alone, inhibits proliferation and increases the G1 compartment in K562 cells. Cell proliferation was evaluated using the MTT cell proliferation assay (top middle). Cell-cycle analysis (representative flow cytometry histograms, bottom) was performed 72 hours after transfection in nocodazole-treated and untreated cells. The bar graph (top right) represents the mean ± SD of 4 independent experiments performed without nocodazole synchronization. Statistical analysis was performed using the Student 2-sided t test: #P < .05; **P < .01.
