Figure 1

Phospholipase D1 (PLD1) is overexpressed in the aggressive, oestrogen receptor (ER)-negative variant of MCF-7 cells, TMX2-28. (A) Cell cultures were grown to subconfluence, and RNA and protein were purified. Oestrogen receptor-α mRNA expression was determined by real-time RT–PCR (bottom), and protein expression was determined by western immunoblotting (top). Experiments were repeated two times; the bottom panel shows the mean and s.d., and the top panel shows a representative image from one experiment. (B) Cell-cycle analysis was determined by propidium iodine staining and FACS analysis. The mean and s.d. from two experiments are shown. The per cent of TMX2-28 cells in each phase of the cell cycle differed significantly from the per cent of MCF-7 cells in the same phase (P<0.05). (C) Total RNA was isolated from cell cultures, and mRNA levels were determined by real-time RT–PCR. Data for each cell line are normalised to hypoxanthine ribosyltransferase (HPRT) and are represented as mean and s.d. from two biological samples each tested in duplicate. Levels of PLD1 mRNA were significantly higher in TMX2-28 cells than in MCF-7, 184, and 184AA2 cells (P<0.05). The same trend was apparent between TMX2-28 and 184A1 cells, but the means were not significantly different (P=0.056).