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Profiling of gene expression changes caused by p53 gain‐of‐function mutant alleles in prostate cancer cells
Author(s) -
Tepper Clifford G.,
Gregg Jeffrey P.,
Shi XuBao,
Vinall Ruth L.,
Baron Colin A.,
Ryan Philip E.,
Desprez PierreYves,
Kung HsingJien,
deVere White Ralph W.
Publication year - 2005
Publication title -
the prostate
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.295
H-Index - 123
eISSN - 1097-0045
pISSN - 0270-4137
DOI - 10.1002/pros.20308
Subject(s) - lncap , biology , gene silencing , gene expression profiling , rna interference , gene expression , gene , mutant , cancer research , prostate cancer , microarray analysis techniques , reporter gene , microbiology and biotechnology , genetics , cancer , rna
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 ( 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‐of‐function, 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. © 2005 Wiley‐Liss, Inc.