Aryl hydrocarbon receptor gene silencing with small inhibitory RNA differentially modulates Ah-responsiveness in MCF-7 and HepG2 cancer cells.

Abstract

Sequence-specific small interfering RNA (siRNA) duplexes can be used for gene silencing in mammalian cells and as mechanistic probes for determining gene function. Transfection of siRNAs for the aryl hydrocarbon receptor (AhR) and AhR nuclear translocator (ARNT) mRNAs in MCF-7 breast cancer cells resulted in a 60 to 80% decrease in levels of AhR and ARNT proteins in whole-cell extracts and decreased binding of nuclear extracts to 32P-labeled dioxin-responsive element. siRNA for the AhR also decreased 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced CYP1A1 protein, CYP1A1-dependent activity, and luciferase activity in cells transfected with an Ah-responsive construct. 17beta-estradiol (E2) induces proliferation of MCF-7 cells through enhanced G0/G1 --> S phase progression, and this response is inhibited in cells cotreated with E2 plus TCDD. The effects of TCDD on E2-induced cell-cycle progress were partially blocked in MCF-7 cells transfected with siRNA for AhR. The results also indicated that siRNA-dependent decreases in AhR protein in MCF-7 cells were accompanied by increased G0/G1 --> S phase progression, suggesting a growth-inhibitory role for the "endogenous" AhR. Surprisingly, TCDD alone induced G0/G1 --> S phase progression and exhibited estrogenic activity in MCF-7 cells transfected with siRNA for the AhR. In contrast, degradation of the AhR in HepG2 liver cancer cells resulted in decreased G0/G1 --> S phase progression, and this was accompanied by decreased expression of cyclin D1, cyclin E, cyclin-dependent kinase 2 (cdk2), and cdk4. In the absence of ligand, the AhR exhibits growth-inhibitory (MCF-7) and growth-promoting (HepG2) activity that is cell context-dependent.

• Authors: Maen Abdelrahim, Roger Smith 3rd, Stephen Safe
• Journal: Molecular pharmacology (Mol Pharmacol) Vol. 63 Issue 6 Pg. 1373-81 (Jun 2003) ISSN: 0026-895X [Print] United States
• PMID: 12761348 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• ARNT protein, human
• DNA-Binding Proteins
• Polychlorinated Dibenzodioxins
• RNA, Small Interfering
• Receptors, Aryl Hydrocarbon
• Teratogens
• Transcription Factors
• Aryl Hydrocarbon Receptor Nuclear Translocator

Topics

• Aryl Hydrocarbon Receptor Nuclear Translocator
• Breast Neoplasms (pathology)
• Cell Cycle (drug effects)
• DNA-Binding Proteins
• Gene Silencing (drug effects)
• Humans
• Lung Neoplasms (pathology)
• Polychlorinated Dibenzodioxins (toxicity)
• RNA, Small Interfering (pharmacology)
• Receptors, Aryl Hydrocarbon (genetics, metabolism)
• Teratogens (toxicity)
• Transcription Factors (metabolism)
• Tumor Cells, Cultured