BACKGROUND. Tumor suppressor p53 mutations are associated with the transition of prostate cancer to metastatic, hormone-refractory disease and stable expression of p53 gain-of-function (p53GOF) alleles support growth of LNCaP in androgen-depleted medium. In this study, we performed gene expression profiling of four LNCaP-p53GOF sublines to test the hypothesis that different p53GOF mutants mediated androgen independence via modulation of a common set of genes. METHODS. Expression profiling was performed using Affymetrix HG-U95Av2 arrays followed by hierarchical clustering to identify expression patterns associated with particular molecular alterations. p53GOF-mediated regulation of Id-1 expression was validated by RT-PCR and dual-lUCiferase reporter assays. RNA interference was used to investigate the effects of Id-1 and Id-3 suppression. RESULTS. LNCaP-p53GOF sublines possessed a molecular signature consisting of 95 differentially regulated genes that could be segregated into two clusters of transcripts indUCed (n = 50) and repressed (n = 45) by p53 GOF expression. To begin validating these genes as effectors of the p53 mutants, we evaluated one of the overexpressed genes, Id-1. RT-PCR confirmed the microarray results and revealed elevated Id-1 levels in LNCaP-p53-P151S (loss-of-function only mutant), thereby implicating p53 mutational inactivation, but not gain-of-function, as a basis for Id-1 deregulation. Reporter assays demonstrated enhanced Id-1 promoter activity in an LNCaP-p53GOF subline. The contribution of Id-1 to p53GOF-mediated biology was demonstrated by the ability of RNAi-mediated gene silencing to decrease both basal and androgen-independent proliferation. CONCLUSIONS. While different p53GOF mutants result in overall distinct expression profiles, they share a common set of differentially-expressed genes that can be used to signify their presence and provide insight into mechanisms underlying androgen independence.
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Tepper, C. G., Gregg, J. P., Shi, X. B., Vinall, R. L., Baron, C. A., Ryan, P. E., Desprez, P. Y., Kung, H. J., & DeVere White, R. W. (2005). Profiling of gene expression changes caused by p53 gain-of-function mutant alleles in prostate cancer cells. Prostate, 65(4), 375-389. https://doi.org/10.1002/pros.20308