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 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.
| Original language | English |
|---|---|
| Pages (from-to) | 375-389 |
| Number of pages | 15 |
| Journal | Prostate |
| Volume | 65 |
| Issue number | 4 |
| DOIs | |
| Publication status | Published - Dec 1 2005 |
| Externally published | Yes |
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Keywords
- Androgen independence
- Id-1
- Microarray
- RNA interference
ASJC Scopus subject areas
- Oncology
- Urology
Cite this
Profiling of gene expression changes caused by p53 gain-of-function mutant alleles in prostate cancer cells. / Tepper, Clifford G.; Gregg, Jeffrey P.; Shi, Xu Bao; Vinall, Ruth L.; Baron, Colin A.; Ryan, Philip E.; Desprez, Pierre Yves; Kung, Hsing Jien; DeVere White, Ralph W.
In: Prostate, Vol. 65, No. 4, 01.12.2005, p. 375-389.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Profiling of gene expression changes caused by p53 gain-of-function mutant alleles in prostate cancer cells
AU - Tepper, Clifford G.
AU - Gregg, Jeffrey P.
AU - Shi, Xu Bao
AU - Vinall, Ruth L.
AU - Baron, Colin A.
AU - Ryan, Philip E.
AU - Desprez, Pierre Yves
AU - Kung, Hsing Jien
AU - DeVere White, Ralph W.
PY - 2005/12/1
Y1 - 2005/12/1
N2 - 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.
AB - 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.
KW - Androgen independence
KW - Id-1
KW - Microarray
KW - RNA interference
UR - http://www.scopus.com/inward/record.url?scp=28744435101&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=28744435101&partnerID=8YFLogxK
U2 - 10.1002/pros.20308
DO - 10.1002/pros.20308
M3 - Article
C2 - 16037992
AN - SCOPUS:28744435101
VL - 65
SP - 375
EP - 389
JO - Prostate
JF - Prostate
SN - 0270-4137
IS - 4
ER -