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Mutual transcriptional interference between RelA and androgen receptor
J J Palvimo1, P Reinikainen, T Ikonen
1Institute of Biomedicine, Department of Physiology, University of Helsinki, FIN-00014 Helsinki, Finland.
Abstract:
Cross-modulation between androgen receptor (AR) and NF-kappaB/Rel proteins was studied using various androgen- and NF-kappaB-regulated reporter genes under transient transfection conditions. In COS-1 cells, elevated expression of RelA (p65) repressed AR-mediated transactivation in a dose-dependent manner, whereas NFkappaB1 (p50), another major member of the NF-kappaB family, did not influence transactivation. The repression of AR appeared to involve the N-terminal region of the protein between residue 297 and the DNA-binding domain. RelA-mediated transrepression could not be overcome by increasing the amount of AR. Transcriptional interference between RelA and AR was mutual in that cotransfected AR was able to attenuate transactivation by RelA in a dose- and steroid-dependent fashion. An excess of RelA was able to rescue the repression to some extent. Immunological analyses of RelA and AR protein levels indicated that transrepression was not due to reciprocal decrease in their amounts. Neither did AR increase the concentration of IkappaBalpha, which can sequester and inactivate RelA. Electrophoretic mobility shift assays using extracts from cotransfected cells and purified recombinant proteins showed that AR and RelA did not significantly influence each other's DNA binding activity. Nevertheless, protein-protein interaction experiments demonstrated a weak association between AR and RelA. Collectively, these data suggest that the mutual repression in intact cells is due to formation of AR-RelA complexes that are held together by another partner or to competition for a coactivator required for transcription.
Insights
Androgen receptor (AR) and NF-kappaB signaling pathways mutually repress each other
Area of Science:
- Molecular Biology
- Cellular Signaling
- Gene Regulation
Background:
- The androgen receptor (AR) and NF-kappaB signaling pathways are crucial in various cellular processes, including development and disease.
- Interactions between these pathways can significantly impact gene expression and cellular function.
Purpose of the Study:
- To investigate the cross-modulation and potential interference between the androgen receptor (AR) and NF-kappaB/Rel proteins.
- To elucidate the molecular mechanisms underlying the observed interactions between AR and NF-kappaB signaling.
Main Methods:
- Utilized reporter gene assays in COS-1 cells to assess AR and NF-kappaB transactivation.
- Performed transient transfections with varying expression levels of AR, RelA (p65), and NFkappaB1 (p50).
- Conducted protein level analyses, electrophoretic mobility shift assays (EMSA), and protein-protein interaction studies.
Main Results:
- Elevated RelA (p65) expression dose-dependently repressed AR-mediated transactivation, involving the AR N-terminal region.
- AR attenuated RelA transactivation in a dose- and steroid-dependent manner, indicating mutual repression.
- Repression was not due to altered protein levels, DNA binding, or IkappaBalpha induction, suggesting complex formation or coactivator competition.
Conclusions:
- AR and NF-kappaB signaling pathways exhibit mutual transcriptional interference.
- The interaction likely involves the formation of AR-RelA complexes or competition for essential coactivators.
- These findings highlight a novel regulatory mechanism influencing cellular responses to androgens and inflammatory signals.