Regulation of u-PA gene expression in human prostate cancer

u-PA contributes to Cap progression, especially in the metastatic androgen- insensitive state. In vitro, u-PA is expressed by androgen-insensitive, but not androgen-sensitive, Cap cell lines. We hypothesized that in androgen-s ensitive Cap an activated ARE represses u-PA expression but In androgen-ins ensitive CaP this repression is lost and u-PA is upregulated through MAP ki nase signaling pathways. To determine whether binding of the DHT-AR complex to AREs in the u-PA promoter region represses u-PA transcription in androg en-sensitive Cap, we studied 2 PC3 androgen-insensitive human CaP cell line s stably transfected with AR [PC3(AR)(2) and PC3(AR)(13)] and I mock-transf ected cell line [PC3(M)]. In the presence of the synthetic androgen miboler one, both PC3(AR)(2) and PC3(AR)(13), but not PC3(M), cells showed decrease d u-PA expression as assayed by Western and Northern blotting. The AR inhib itor flutamide abrogated mibolerone's effect. Androgen regulation of a seco nd gene, PSA, was also demonstrated in the PC3(AR)(2) cell line. To explore the pathway stimulating u-PA expression in Cap, we performed transient tra nsfections in PC3(AR)(2) cells using u-PA promoter-regulated CAT reporter c onstructs. Compared to full-length u-PA promoter-CAT constructs, either del etion or mutation of the 5' AP-I or PEA3 site reduced CAT expression. The l ocation of androgen responsiveness in the u-PA promoter was not identified through the combination of promoter search and transient transfection assay s, indicating that a more complicated mechanism is involved in the AR-media ted downmodulation of u-PA expression. (C) 2001 Wiley-Liss, Inc.