title: Profiling of gene expression changes caused by p53 gain-of-function mutant alleles in prostate cancer cells.
abstract: 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 p53GOF 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.
authors.affiliation: Department of Biochemistry and Molecular Medicine, University of California, Davis School of Medicine, Division of Basic Sciences, UC Davis Cancer Center, Sacramento, California 95817, USA. cgtepper@ucdavis.edu
authors.forename: Clifford G
authors.initials: CG
authors.lastname: Tepper
authors.forename: Jeffrey P
authors.initials: JP
authors.lastname: Gregg
authors.forename: Xu-Bao
authors.initials: XB
authors.lastname: Shi
authors.forename: Ruth L
authors.initials: RL
authors.lastname: Vinall
authors.forename: Colin A
authors.initials: CA
authors.lastname: Baron
authors.forename: Philip E
authors.initials: PE
authors.lastname: Ryan
authors.forename: Pierre-Yves
authors.initials: PY
authors.lastname: Desprez
authors.forename: Hsing-Jien
authors.initials: HJ
authors.lastname: Kung
authors.forename: Ralph W
authors.initials: RW
authors.lastname: deVere White
mesh_terms: D000483:Alleles; D045744:Cell Line, Tumor; D020869:Gene Expression Profiling; D016158:Genes, p53 / Q000235:genetics*; D006801:Humans; D051798:Inhibitor of Differentiation Protein 1 / Q000096:biosynthesis / Q000235:genetics; D008297:Male; D009154:Mutation; D009376:Neoplasms, Hormone-Dependent / Q000235:genetics* / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D011401:Promoter Regions, Genetic; D011471:Prostatic Neoplasms / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D012334:RNA, Neoplasm / Q000737:chemistry / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D051798:Inhibitor of Differentiation Protein 1; D012334:RNA, Neoplasm; D034741:RNA, Small Interfering