期刊
PROSTATE
卷 65, 期 4, 页码 375-389出版社
WILEY
DOI: 10.1002/pros.20308
关键词
microarray; androgen independence; Id-1; RNA interference
资金
- NCI NIH HHS [R01-CA77662, P30 CA93373-01] Funding Source: Medline
BACKGROUND: Tumor suppressor p53 mutations are associated with the transition of Prostate cancer to metastatic, hormone-refractory disease and stable expression of p53 gain-offunction (P53(GOF)) alleles support growth of LNCaP in androgen-depleted medium. In this study, we performed gene expression profiling of four LNCaP-p53(GOF) sublines to test the hypothesis that different p53(GOF) 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. p53(GOF)-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-p53(GOF) 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-offunction, as a basis for Id-1 deregulation. Reporter assays demonstrated enhanced Id-1 promoter activity in an LNCaP-p53(GOF) subline. The contribution of Id-1 to p53(GOF)-mediated biology was demonstrated by the ability of RNAi-mediated gene silencing to decrease both basal and androgen-independent proliferation. CONCLUSIONS: While different p53(GOF) 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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