Dominance of the α_1B-Adrenergic Receptor and its Subcellular Localization in Human and TRAMP Prostate Cancer Cell Lines

Abstract

The function and distribution of α₁-adrenergic receptor (AR) subtypes in prostate cancer cells is well characterized. Previous studies have used RNA localization or low-avidity antibodies in tissue or cell lines to determine the α₁-AR subtype and suggested that the α_{1 A}-AR is dominant. Two androgen-insensitive, human metastatic cancer cell lines DU145 and PC3 were used as well as the mouse TRAMP C1-C3 primary and clonal cell lines. The density of α₁-ARs was determined by saturation binding and the distribution of the different α₁-AR subtypes was examined by competition-binding experiments. In contrast to previous studies, the major α₁-AR subtype in DU145, PC3 and all of the TRAMP cell lines is the α_1B-AR. DU145 cells contained 100% of the α_1B-AR subtype, whereas PC3 cells were composed of 21% α_{1 A}-AR and 79% α_1B-AR. TRAMP cell lines contained between 66% and 79% of the α_1B-AR with minor fractions of the other two subtypes. Faster doubling time in the TRAMP cell lines correlated with decreasing α _1B-AR and increasing α_{1 A}- and α_1D-AR densities. Transfection with EGFP-tagged α_1B-ARs revealed that localization was mainly intracellular, but the majority of the receptors translocated to the cell surface after extended preincubation (18 hr) with either agonist or antagonist. Localization was confirmed by ligand-binding studies and inositol phosphate assays where prolonged preincubation with either agonist and/or antagonist increased the density and function of α ₁-ARs, suggesting that the native receptors were mostly intracellular and nonfunctional. Our studies indicate that α_1B-ARs are the major α₁-AR subtype expressed in DU145, PC3, and all TRAMP cell lines, but most of the receptor is localized in intracellular compartments in a nonfunctional state, which can be rescued upon prolonged incubation with any ligand.

Key Words: :

• Prostate cancer
• Adrenergic receptor
• Intracellular localization
• DU145
• PC3
• TRAMP.