Coordinate regulation of bovine prion protein gene promoter activity by two Sp1 binding site polymorphisms

Relationships between insertion/deletion (Ins/Del) polymorphisms of the bovine prion protein gene (PRNP) promoter and bovine spongiform encephalopathy (BSE) susceptibility have been reported. Our previous study has shown that polymorphisms of −6C → T included in the specific protein 1 (Sp1) site in the 5′-flanking region of bovine PRNP influence the promoter activity of bovine PRNP. The present study shows that 12 and 23 bp Ins/Del polymorphisms in the upstream region and an additional polymorphism (−47C → A) in the Sp1 binding site coordinately affect the promoter activity. Reporter gene assays demonstrated that the bovine PRNP promoter containing −47A and 23 bp Del/12 bp Ins or 23 bp Ins/12 bp Ins showed lower promoter activity compared with other haplotypes (23 bp Del/12 bp Ins or 23 bp Ins/12 bp Del with −47C) or the wild-type haplotype (23 bp Del/12 bp Del with −47C). Furthermore, gel shift assays showed that the binding activity of Sp1 to the PRNP promoter was influenced by both polymorphisms with corresponding effects on the promoter activity. The coordinate regulation of the bovine PRNP promoter suggests the two Sp1 binding site polymorphisms control Sp1 binding to the PRNP promoter and its activity.     

Secretoglobin 2A1 is under selective androgen control mediated by a peculiar binding site for Sp family transcription factors

Human secretoglobin (SCGB) 2A1 (or lipophilin C, lacryglobin, mammaglobin B) is a small protein of unknown function that forms heterodimers with secretoglobin 1D1 (lipophilin A) in tears and is expressed in the prostate. Here we show that SCGB 2A1 is under androgen control in the androgen-responsive prostatic cell line LNCaP and can be induced more than 20-fold by dihydrotestosterone. Only 6 h after androgen treatment, a strong DNase I-hypersensitive site is induced in the proximal promoter within chromatin. Within the boundaries of this DNase I-hypersensitive site a minimal 32-bp peculiar dimeric inverted repeat variant GC box (dim-IR-GA box) was found to confer androgen but not glucocorticoid responsiveness in gene transfer experiments. Mutations of both GA boxes that abolish binding of Sp1 and Sp3 also abrogate the androgen response. In an EMSA the DNA binding domain of the androgen receptor (AR) was not able to bind directly to the dim-IR-GA box. However, AR is functionally required for the hormone response because induction can be inhibited with the nonsteroidal antagonist bicalutamide. Chromatin immunoprecipitation experiments demonstrated that AR is recruited to the proximal promoter 10 min after androgen treatment. Therefore we propose that SCGB 2A1 represents a new class of androgen target genes that are purely under indirect AR control mediated by DNA-bound Sp factors.