The interleukin 6 receptor is a direct transcriptional target of E2F3 in prostate tumor derived cells

Stephen J. Libertini, Honglin Chen, Bushra Al-Bataina, Tilak Koilvaram, Michael George, Allen C Gao, Maria Mudryj

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

BACKGROUND The E2F/RB pathway is frequently disrupted in multiple human cancers. E2F3 levels are elevated in prostate tumors and E2F3 overexpression independently predicts clinical outcome. The goals of this study were to identify direct transcriptional targets of E2F3 in prostate tumor derived cells. METHODS Expression array studies identified the interleukin 6 receptor (IL-6R) as an E2F3 target. E2F3-dependent expression of IL-6R was analyzed by real time PCR and Western immunoblot analysis in several cell lines. Chromatin immunoprecipitation (ChIP) and IL-6R-luciferase reporter plasmid studies were used to characterize the IL-6R promoter. RESULTS Expression array studies identified genes that were regulated by E2F3 in prostate tumor derived cell lines. The network most significantly associated with E2F3-regulated transcripts was cytokine signaling and the IL-6R was a component of several of the most prominent E2F3-regulated pathways. The transcriptional regulation of IL-6R by E2F3 knockdown was validated in several prostate tumor-derived cell lines at the RNA level and protein level. The IL-6R regulatory region containing ChIP-identified E2F3 binding sites was cloned into a reporter and co-transfected with an E2F3a expression plasmid. The luciferase assay showed that E2F3a transactivated the IL-6R promoter in a dose dependent manner. The functional consequence of IL-6R decrease was a reduction in the levels of ERK1/2 phosphorylation, indicating that IL-6R initiated signaling was altered. CONCLUSION These studies connect the E2F and IL-6 signaling cascade, thus providing the mechanistic link between two major regulatory networks that are perturbed during prostate tumorigenesis.

Original languageEnglish (US)
Pages (from-to)649-660
Number of pages12
JournalProstate
Volume72
Issue number6
DOIs
StatePublished - May 1 2012

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Interleukin-6 Receptors
Prostate
Neoplasms
Chromatin Immunoprecipitation
Tumor Cell Line
Luciferases
Plasmids
Nucleic Acid Regulatory Sequences
Real-Time Polymerase Chain Reaction
Interleukin-6
Carcinogenesis
Western Blotting
Binding Sites
Phosphorylation
RNA
Cytokines

Keywords

  • chromatin immunoprecipitation
  • cytokine
  • promoter
  • regulation

ASJC Scopus subject areas

  • Urology
  • Oncology

Cite this

The interleukin 6 receptor is a direct transcriptional target of E2F3 in prostate tumor derived cells. / Libertini, Stephen J.; Chen, Honglin; Al-Bataina, Bushra; Koilvaram, Tilak; George, Michael; Gao, Allen C; Mudryj, Maria.

In: Prostate, Vol. 72, No. 6, 01.05.2012, p. 649-660.

Research output: Contribution to journalArticle

Libertini, Stephen J. ; Chen, Honglin ; Al-Bataina, Bushra ; Koilvaram, Tilak ; George, Michael ; Gao, Allen C ; Mudryj, Maria. / The interleukin 6 receptor is a direct transcriptional target of E2F3 in prostate tumor derived cells. In: Prostate. 2012 ; Vol. 72, No. 6. pp. 649-660.
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AU - Libertini, Stephen J.

AU - Chen, Honglin

AU - Al-Bataina, Bushra

AU - Koilvaram, Tilak

AU - George, Michael

AU - Gao, Allen C

AU - Mudryj, Maria

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N2 - BACKGROUND The E2F/RB pathway is frequently disrupted in multiple human cancers. E2F3 levels are elevated in prostate tumors and E2F3 overexpression independently predicts clinical outcome. The goals of this study were to identify direct transcriptional targets of E2F3 in prostate tumor derived cells. METHODS Expression array studies identified the interleukin 6 receptor (IL-6R) as an E2F3 target. E2F3-dependent expression of IL-6R was analyzed by real time PCR and Western immunoblot analysis in several cell lines. Chromatin immunoprecipitation (ChIP) and IL-6R-luciferase reporter plasmid studies were used to characterize the IL-6R promoter. RESULTS Expression array studies identified genes that were regulated by E2F3 in prostate tumor derived cell lines. The network most significantly associated with E2F3-regulated transcripts was cytokine signaling and the IL-6R was a component of several of the most prominent E2F3-regulated pathways. The transcriptional regulation of IL-6R by E2F3 knockdown was validated in several prostate tumor-derived cell lines at the RNA level and protein level. The IL-6R regulatory region containing ChIP-identified E2F3 binding sites was cloned into a reporter and co-transfected with an E2F3a expression plasmid. The luciferase assay showed that E2F3a transactivated the IL-6R promoter in a dose dependent manner. The functional consequence of IL-6R decrease was a reduction in the levels of ERK1/2 phosphorylation, indicating that IL-6R initiated signaling was altered. CONCLUSION These studies connect the E2F and IL-6 signaling cascade, thus providing the mechanistic link between two major regulatory networks that are perturbed during prostate tumorigenesis.

AB - BACKGROUND The E2F/RB pathway is frequently disrupted in multiple human cancers. E2F3 levels are elevated in prostate tumors and E2F3 overexpression independently predicts clinical outcome. The goals of this study were to identify direct transcriptional targets of E2F3 in prostate tumor derived cells. METHODS Expression array studies identified the interleukin 6 receptor (IL-6R) as an E2F3 target. E2F3-dependent expression of IL-6R was analyzed by real time PCR and Western immunoblot analysis in several cell lines. Chromatin immunoprecipitation (ChIP) and IL-6R-luciferase reporter plasmid studies were used to characterize the IL-6R promoter. RESULTS Expression array studies identified genes that were regulated by E2F3 in prostate tumor derived cell lines. The network most significantly associated with E2F3-regulated transcripts was cytokine signaling and the IL-6R was a component of several of the most prominent E2F3-regulated pathways. The transcriptional regulation of IL-6R by E2F3 knockdown was validated in several prostate tumor-derived cell lines at the RNA level and protein level. The IL-6R regulatory region containing ChIP-identified E2F3 binding sites was cloned into a reporter and co-transfected with an E2F3a expression plasmid. The luciferase assay showed that E2F3a transactivated the IL-6R promoter in a dose dependent manner. The functional consequence of IL-6R decrease was a reduction in the levels of ERK1/2 phosphorylation, indicating that IL-6R initiated signaling was altered. CONCLUSION These studies connect the E2F and IL-6 signaling cascade, thus providing the mechanistic link between two major regulatory networks that are perturbed during prostate tumorigenesis.

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