Figure 1
PRL-3 expression positively correlates with mTOR activity in vivo and in vitro.
(a) Immunoblotting of matched pairs of tumour-normal tissues from PRL-3-positive cancer patients. The ratio of phosphorylated/total 4E-BP1 band densities were calculated and normalized to the phosphorylated/total 4E-BP1 ratio in lane 1. Asterisks, patient tumour samples wherein 4E-BP1 hyperphosphorylation correlates with PRL-3 expression. (b) Immunoblotting of normal and MMTV-PyMT mammary tissues over the course of spontaneous tumour development. Top panel, images of normal mice (N) or transgenic MMTV-PyMT mice (T) between the ages of 6 to 15 weeks. Middle panel, representative images of excised breast tissues from mice. All images were photographed by Thura Min. Blue dashed lines, gross size of palpable mammary tumours in MMTV-PyMT mice. Lower panels, correlation between PRL-3 expression and phosphorylation of 4E-BP1 and mTOR. (c–d) Overexpression of EGFP vector only (Vec), EGFP-tagged wild-type PRL-3 (PRL-3) or EGFP-tagged catalytic-inactive PRL-3 (PRL-3 C104S) in a panel of 9 human cancer cell lines. PRL-3 upregulates mTOR phosphorylation in (c) HCT116, HeLa, MCF7, HCT15, LOVO and SW620 cells, but not in (d) G361, DLD1 or A2780 cells. GAPDH served as a loading control. The ratio of phosphorylated/total mTOR band densities were calculated and normalized to the phosphorylated/total mTOR ratio in lane 1. Full immunoblots for the cropped images presented here are provided in the Supplementary Information.
