Sp1 and Sp3 transcription factors upregulate the proximal promoter of the human prostate-specific antigen gene in prostate cancer cells

The serum level of prostate-specific antigen (PSA) is useful as a clinical marker for diagnosis and assessment of the progression of prostate cancer, and in evaluating the effectiveness of treatment. We characterized four Sp1/Sp3 binding sites in the proximal promoter of the PSA gene. In a luciferase assay, these sites contributed to the basal promoter activity in prostate cancer cells. In an electrophoretic mobility shift assay and chromatin immunoprecipitation assay, we confirmed that Sp1 and Sp3 bind to these sites. Overexpression of wild-type Sp1 and Sp3 further upregulated the promoter activity, whereas overexpression of the Sp1 dominant-negative form or addition of mithramycin A significantly reduced the promoter activity and the endogenous mRNA level of PSA. Among the four binding sites, a GC box located at nucleotides -53 to -48 was especially critical for basal promoter activity. These results indicate that Sp1 and Sp3 are involved in the basal expression of PSA in prostate cancer cells.     

Induction of dystrophin Dp71 expression during neuronal differentiation: opposite roles of Sp1 and AP2α in Dp71 promoter activity

In this study, we delineated the molecular mechanisms that modulate Dp71 expression during neuronal differentiation, using the N1E‐115 cell line. We demonstrated that Dp71 expression is up‐regulated in response to cAMP‐mediated neuronal differentiation of these cells, and that this induction is controlled at promoter level. Functional deletion analysis of the Dp71 promoter revealed that a 5′‐flanking 159‐bp DNA fragment that contains Sp1 and AP2 binding sites is necessary and sufficient for basal expression of this TATA‐less promoter, as well as for its induction during neuronal differentiation. Electrophoretic mobility shift and chromatin immunoprecipitation assays revealed that Sp1 and AP2α bind to their respective DNA elements within the Dp71 basal promoter. Overall, mutagenesis assays on the Sp1 and AP2 binding sites, over‐expression of Sp1 and AP2α, as well as knock‐down experiments on Sp1 and AP2α gene expression established that Dp71 basal expression is controlled by the combined action of Sp1 and AP2α, which act as activator and repressor, respectively. Furthermore, we demonstrated that induction of Dp71 expression in differentiated cells is the result of the maintenance of positive regulation exerted by Sp1, as well as of the loss of AP2α binding, which ultimately releases the promoter from repression.