Figure 3.

Specificity of AR:ART-27 interactions. Interaction of ART-27 with the AR N terminus (N-T; rat AR_18–500), AR LBD (rat AR_579–901), and other transcriptional regulatory factors was analyzed using the modified yeast two-hybrid assay. Regulatory factors examined include the GR AF-1 (rat GR_107–237), GR 30IIB (rat GR_107–237 E219K, F220L, W234R), SRC-1 (SRC-1_374–800), GR LBD (rat GR_525–795), TAF_II130 (TAF_II130_270–700), Sp1 A (Sp1_83–262), Sp1 B (Sp1_263–524), CREB-N (CREB_3–296), ERα(ERα_1–595), and VP16. The strength of interaction was determined by a quantitative liquid β-galactosidase assay after a 24-h incubation in galactose medium at 30°C and normalized to protein expression using the common HA epitope resident on each protein. The background from the vector-only control was subtracted from each sample. The interaction with ERα was observed in the presence, but not the absence (Markus, Taneja, Logan, Li, Ha, Hittelman, Rogatsky, and Garabedian, unpublished results), of 100 nM 17β-estradiol. No interaction was observed with AR LBD and the GR LBD in either the presence or absence of 100 nM R1881 and 10 μM deoxycorticosterone, the preferred GR ligand in yeast (Garabedian and Yamamoto, 1992).