Figure 3.
siRNA knockdown experiments. A: Reverse transcription-polymerase chain reaction (RT-PCR) analysis. Two CTCL cell lines HUT78 and MyLa were transfected with 2.5 μg of control, FRA2, JUNB, or JUND siRNA. After 48 h, total RNAs were extracted. RT-PCR was performed for the transcripts of FRA2, JUNB, JUND, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). The representative results from three separate experiments are shown. B: Cell growth analysis. HUT78, MyLa, and HSB-2 (a control T-ALL cell line) were transfected with 2.5 μg of control, FRA2, JUNB, or JUND siRNA, and cultured in a 96-well plate at 0.5×104 cells/well. At the indicated time points, viable cell numbers were determined on a FACSCalibur instrument by gating out cells stained with propidium iodide. Data are shown as the mean±SEM from three separate experiments. *p<0.05. C: Double knockdown experiments. HUT78 and MyLa were transfected with siRNAs for the control, FRA2, or JUND as indicated, and cultured in a 96-well plate at 0.5×104 cells/well. After 4 days, viable cell numbers were determined on a FACSCalibur instrument by gating out cells stained with propidium iodide. Data are shown as the mean±SEM from three separate experiments. *p<0.05. D: Quantitation of transcripts. Real-time quantitative PCR was performed to quantitate the transcripts of CCR4, MYB, MDM2, and BCL6 relative to β2-microglobulin used as an internal control. Data are shown as the mean±SEM from three separate experiments. *p<0.05.