Fig 3.

EGF does not alter the Aurora-A mRNA level in human colorectal cancer cell lines. (A) Serum-starved LS174T cells were treated with EGF for 3 hrs, and total RNA was harvested. The expression levels of Aurora-A and GAPDH mRNA were detected by RT-PCR. The lower panel, serum-starved cells were treated with EGF for different time-points, and the expression level of Aurora-A mRNA was quantified by real-time RT-PCR and normalized with RPL13A (ribosomal protein L 13A). The mean ± S.D. (error bars) was obtained from three independent experiments. (B) Neither U0126 nor wortmannin changed the expression level of Aurora A mRNA under EGF treatment. The expression of Aurora-A mRNA was analysed as in (A). (C) Cells were transiently transfected with the Aurora-A promoter-luciferase construct. After starvation, cells were treated with EGF for 0, 4, 18 and 24 hrs, and the reporter assay was performed with a dual luciferase assay kit. The mean ± S.D. was obtained from three independent experiments. (D) Serum-starved cells were pre-treated with actinomycin D for 30 min., followed by EGF for another 2 hrs. The expressions of Aurora-A and α-tubulin were analysed by Western Blot.