Figure 1

1a. RNA response to exogenous CT at 48 h is calcitonin (CT) receptor-dependent. CD44 total and variant RNA are quantified by triplicate TaqMan RT-PCR experiments. In benign BPH-1 cells, no response is observed to exogenous calcitonin (CT). CT-cells, a PC-3M derivative that lack endogenous CT but have CT receptor, showed a decrease in CD44 total RNA but an increase in variant RNA. The G_sα-QL cell derivative showed the same trend. In CT-receptor-negative CTR- and PC-3 cells, there is no significant response to CT. Error bars are standard deviation. *p = 0.01; ^† p = 0.05 with respect to mock treated controls. 1b. Western blot analysis. Exogenous calcitonin given to the CT-cells for 3 h stimulates expression of CD44s at 75 kD (top), and CD44v at 180 kD and its cleavage products below 97 kD. Comparison is shown to cells with no treatment (NT). β-actin analysis control confirms equal protein loading (bottom). 1c. This stimulatory effect persists at 48 h and at doses of 50 mM or 250 mM exogenous calcitonin. Also, a time-dependent reduction in protein was observed after cycloheximide treatment, suggesting that CT effect on CD44 variant requires de novo protein synthesis and does not result from protein stabilization. 1d. CT+ cells, endogenously expressing CT, are derived from PC-3M cells and have increased RNA for CD44v (p = 0.02) but a decrease in CD44 total (p = 0.008).